Scientists reveal surprising picture of how powerful antibody neutralizes HIV by Biomechanism

“The findings advance AIDS vaccine development”

Researchers at The Scripps Research Institute have uncovered the surprising details of how a powerful anti-HIV antibody grabs hold of the virus. The findings, published in Science Express on October 13, 2011, highlight a major vulnerability of HIV and suggest a new target for vaccine development.

Caption: This is the PGT 128 antibody in action. Credit: Wilson lab, The Scripps Research Institute

“What’s unexpected and unique about this antibody is that it not only attaches to the sugar coating of the virus but also reaches through to grab part of the virus’s envelope protein,” said the report’s co-senior author Dennis Burton, a professor at The Scripps Research Institute and scientific director of the International AIDS Vaccine Initiative’s (IAVI) Neutralizing Antibody Center, based on the Scripps Research La Jolla campus.

“We can now start to think about constructing mimics of these viral structures to use in candidate vaccines,” said co-senior author Ian Wilson, who is Hansen Professor of Structural Biology and member of the Skaggs Institute for Chemical Biology at Scripps Research.

Other institutions in the United States, United Kingdom, Japan, and the Netherlands contributed to the research as part of an ongoing global HIV vaccine development effort.

Getting a Better Grip on HIV

Researchers from the current team recently isolated the new antibody and 16 others from the blood of HIV-infected volunteers, in work they reported online in the journal Nature on August 17, 2011. Since the 1990s, Burton, Wilson, and other researchers have been searching for such “broadly neutralizing” antibodies against HIV—antibodies that work against many of the various strains of the fast-mutating virus—and by now have found more than a dozen. PGT 128, the antibody described in the new report, can neutralize about 70 percent of globally circulating HIV strains by blocking their ability to infect cells. It also can do so much more potently—in other words, in smaller concentrations of antibody molecules—than any previously reported broadly neutralizing anti-HIV antibody.

The new report illuminates why PGT 128 is so effective at neutralizing HIV. Using the Wilson lab’s expertise in X-ray crystallography, Robert Pejchal, a research associate in the Wilson lab, determined the structure of PGT 128 joined to its binding site on molecular mockups of the virus, designed in part by Robyn Stanfield and Pejchal in the Wilson group and Bill Schief, now an IAVI principal scientist and associate professor at Scripps Research, and his group. With these structural data, and by experimentally mutating and altering the viral target site, they could see that PGT 128 works in part by binding to glycans on the viral surface.

Thickets of these sugars normally surround HIV’s envelope protein, gp120, largely shielding it from attack by the immune system. Nevertheless, PGT 128 manages to bind to two closely spaced glycans, and at the same time reaches through the rest of the “glycan shield” to take hold of a small part of structure on gp120 known as the V3 loop. This penetration of the glycan shield by PGT 128 was also visualized by electron microscopy with a trimeric form of the gp120/gp41 envelope protein of HIV-1 by Reza Kayat and Andrew Ward of Scripps Research; this revealed that the PGT 128 epitope appears to be readily accessible on the virus.

“Both of these glycans appear in most HIV strains, which helps explain why PGT 128 is so broadly neutralizing,” said Katie J. Doores, a research associate in the Burton lab who was one of the report’s lead authors. PGT 128 also engages V3 by its backbone structure, which doesn’t vary as much as other parts of the virus because it is required for infection.

PGT 128′s extreme potency is harder to explain. The antibody binds to gp120 in a way that presumably disrupts its ability to lock onto human cells and infect them. Yet it doesn’t bind to gp120 many times more tightly than other anti-HIV antibodies. The team’s analysis hints that PGT 128 may be extraordinarily potent because it also binds two separate gp120 molecules, thus tying up not one but two cell-infecting structures. Other mechanisms may also be at work.

Toward an AIDS Vaccine

Researchers hope to use the knowledge of these antibodies’ binding sites on HIV to develop vaccines that stimulate a long-term—perhaps lifetime—protective antibody response against those same vulnerable sites.

“We’ll probably need multiple targets on the virus for a successful vaccine, but certainly PGT 128 shows us a very good target,” said Burton.

Intriguingly, the basic motif of PGT 128′s target may mark a general vulnerability for HIV. “Other research is also starting to suggest that you can grab onto two glycans and a beta strand and get very potent and broad neutralizing antibodies against HIV,” Wilson said.

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The research was supported by the International AIDS Vaccine Initiative, National Institutes of Health, the U.S. Department of Energy, the Canadian Institutes of Health Research, the UK Research Councils, the Ragon Institute, and other organizations.

From Blue Whales to Earthworms, a Common Mechanism Gives Shape to Living Beings

This is a diagram of the mechanism for form. The blue whale and the earth worm owe their form to the same biological mechanism. EPFL researchers have discovered the secret. (Credit: Infographic courtesy of Pascal Coderay, EPFL)

Science Daily  — Why don't our arms grow from the middle of our bodies? The question isn't as trivial as it appears. Vertebrae, limbs, ribs, tailbone ... in only two days, all these elements take their place in the embryo, in the right spot and with the precision of a Swiss watch. Intrigued by the extraordinary reliability of this mechanism, biologists have long wondered how it works. Now, researchers at EPFL (Ecole Polytechnique Fédérale de Lausanne) and the University of Geneva (Unige) have solved the mystery.

The embryo is built one layer at a time

Their discovery will be published October 13, 2011 in the journalScience.

During the development of an embryo, everything happens at a specific moment. In about 48 hours, it will grow from the top to the bottom, one slice at a time -- scientists call this the embryo's segmentation. "We're made up of thirty-odd horizontal slices," explains Denis Duboule, a professor at EPFL and Unige. "These slices correspond more or less to the number of vertebrae we have."

Every hour and a half, a new segment is built. The genes corresponding to the cervical vertebrae, the thoracic vertebrae, the lumbar vertebrae and the tailbone become activated at exactly the right moment one after another. "If the timing is not followed to the letter, you'll end up with ribs coming off your lumbar vertebrae," jokes Duboule. How do the genes know how to launch themselves into action in such a perfectly synchronized manner? "We assumed that the DNA played the role of a kind of clock. But we didn't understand how."

When DNA acts like a mechanical clock

Very specific genes, known as "Hox," are involved in this process. Responsible for the formation of limbs and the spinal column, they have a remarkable characteristic. "Hox genes are situated one exactly after the other on the DNA strand, in four groups. First the neck, then the thorax, then the lumbar, and so on," explains Duboule. "This unique arrangement inevitably had to play a role."

The process is astonishingly simple. In the embryo's first moments, the Hox genes are dormant, packaged like a spool of wound yarn on the DNA. When the time is right, the strand begins to unwind. When the embryo begins to form the upper levels, the genes encoding the formation of cervical vertebrae come off the spool and become activated. Then it is the thoracic vertebrae's turn, and so on down to the tailbone. The DNA strand acts a bit like an old-fashioned computer punchcard, delivering specific instructions as it progressively goes through the machine.

"A new gene comes out of the spool every ninety minutes, which corresponds to the time needed for a new layer of the embryo to be built," explains Duboule. "It takes two days for the strand to completely unwind; this is the same time that's needed for all the layers of the embryo to be completed."

This system is the first "mechanical" clock ever discovered in genetics. And it explains why the system is so remarkably precise.

This discovery is the result of many years of work. Under the direction of Duboule and Daniël Noordermeer, the team analyzed thousands of Hox gene spools. With assistance from the Swiss Institute for Bioinformatics, the scientists were able to compile huge quantities of data and model the structure of the spool and how it unwinds over time.

The snake: a veritable vertebral assembly line

The process discovered at EPFL is shared by numerous living beings, from humans to some kinds of worms, from blue whales to insects. The structure of all these animals -- the distribution of their vertebrae, limbs and other appendices along their bodies -- is programmed like a sheet of player-piano music by the sequence of Hox genes along the DNA strand.

The sinuous body of the snake is a perfect illustration. A few years ago, Duboule discovered in these animals a defect in the Hox gene that normally stops the vertebrae-making process.

"Now we know what's happening. The process doesn't stop, and the snake embryo just keeps on making vertebrae, all identical, until the process just runs out of steam."

The Hox clock is a demonstration of the extraordinary complexity of evolution. One notable property of the mechanism is its extreme stability, explains Duboule. "Circadian or menstrual clocks involve complex chemistry. They can thus adapt to changing contexts, but in a general sense are fairly imprecise. The mechanism that we have discovered must be infinitely more stable and precise. Even the smallest change would end up leading to the emergence of a new species."

Schizophrenia Genetics Linked to Disruption in How Brain Processes Sound

Staining performed by Konrad Talbot, PhD, targeting a marker for nerve cells involved in inhibition are shown in cross sections of the hippocampus, which is a part of the brain known to be affected in schizophrenia and involved in memory and cognition. In normal mice (top; A and B) a number of inhibitory cells are found. This staining is reduced in mice with reduced dysbindin (bottom; C and D). The finding is identical to that found in tissue from schizophrenia patients and supports the functional finding of the paper that fast inhibitory processes are disrupted in schizophrenia, leading to symptoms of the disease. (Credit: Konrad Talbot, PhD, Perelman School of Medicine, University of Pennsylvania, Neuron)

Science Daily  — Recent studies have identified many genes that may put people with schizophrenia at risk for the disease. But, what links genetic differences to changes in altered brain activity in schizophrenia is not clear. Now, three labs at the Perelman School of Medicine at the University of Pennsylvania have come together using electrophysiological, anatomical, and immunohistochemical approaches -- along with a unique high-speed imaging technique -- to understand how schizophrenia works at the cellular level, especially in identifying how changes in the interaction between different types of nerve cells leads to symptoms of the disease.

"Our work provides a model linking genetic risk factors for schizophrenia to a functional disruption in how the brain responds to sound, by identifying reduced activity in special nerve cells that are designed to make other cells in the brain work together at a very fast pace" explains lead author Gregory Carlson, PhD, assistant professor of Neuroscience in Psychiatry. "We know that in schizophrenia this ability is reduced, and now, knowing more about why this happens may help explain how loss of a protein called dysbindin leads to some symptoms of schizophrenia."

The findings are reported this week in the Proceedings of the National Academy of Sciences.

Previous genetic studies had found that some forms of the gene for dysbindin were found in people with schizophrenia. Most importantly, a prior finding at Penn showed that the dysbindin protein is reduced in a majority of schizophrenia patients, suggesting it is involved in a common cause of the disease.

For the current PNAS study, Carlson, Steven J. Siegel, MD, PhD, associate professor of Psychiatry, director of the Translational Neuroscience Program; and Steven E. Arnold, MD, director of the Penn Memory Center, used a mouse with a mutated dysbindin gene to understand how reduced dysbindin protein may cause symptoms of schizophrenia.

The team demonstrated a number of sound-processing deficits in the brains of mice with the mutated gene. They discovered how a specific set of nerve cells that control fast brain activity lose their effectiveness when dysbindin protein levels are reduced. These specific nerve cells inhibit activity, but do so in an extremely fast pace, essentially turning large numbers of cells on and off very quickly in a way that is necessary to normally process the large amount of information travelling into and around the brain.

Other previous work at Penn in the lab of Michael Kahana, PhD has shown that in humans the fast brain activity that is disrupted in mice with the dysbindin mutation is also important for short-term memory. This type of brain activity is reduced in people with schizophrenia and resistant to current therapy. Taken as a whole, this work may suggest new avenues of treatment for currently untreatable symptoms of schizophrenia, says Carlson.

Additional co-authors are: Konrad Talbot, Tobias B. Halene, Michael J. Gandal, Hala A. Kazi, Laura Schlosser, Quan H. Phung, and Raquel E. Gur, all from the Department of Psychiatry at Penn.

This work was funded in part by the National Institutes of Mental Health.

Carbon Nanotube Muscles Generate Giant Twist for Novel Motors

This is a scanning electron micrograph image of a 3.8-micron diameter carbon nanotube yarn that functions as a torsional muscle when filled with an ionically conducting liquid and electrochemically charged. The angle ± indicates the deviation between nanotube orientation and yarn direction for this helical yarn. (Credit: Image courtesy of the University of Texas at Dallas)

Science Daily  — New artificial muscles that twist like the trunk of an elephant, but provide a thousand times higher rotation per length, have been developed by a team of researchers from The University of Texas at Dallas, The University of Wollongong in Australia, The University of British Columbia in Canada, and Hanyang University in Korea.

The research appears in the journalScience.

These muscles, based on carbon nanotubes yarns, accelerate a 2000 times heavier paddle up to 590 revolutions per minute in 1.2 seconds, and then reverse this rotation when the applied voltage is changed. The demonstrated rotation of 250 per millimeter of muscle length is over a thousand times that of previous artificial muscles, which are based on ferroelectrics, shape memory alloys, or conducting organic polymers. The output power per yarn weight is comparable to that for large electric motors, and the weight-normalized performance of these conventional electric motors severely degrades when they are downsized to millimeter scale.

These muscles exploit strong, tough, highly flexible yarns of carbon nanotubes, which consist of nanoscale cylinders of carbon that are ten thousand times smaller in diameter than a human hair. Important for success, these nanotubes are spun into helical yarns, which means that they have left and right handed versions (like our hands), depending upon the direction of rotation during twisting the nanotubes to make yarn. Rotation is torsional, meaning that twist occurs in one direction until a limiting rotation results, and then rotation can be reversed by changing the applied voltage. Left and right hand yarns rotate in opposite directions when electrically charged, but in both cases the effect of charging is to partially untwist the yarn.

Unlike conventional motors, whose complexity makes them difficult to miniaturize, the torsional carbon nanotube muscles are simple to inexpensively construct in either very long or millimeter lengths. The nanotube torsional motors consist of a yarn electrode and a counter-electrode, which are immersed in an ionically conducting liquid. A low voltage battery can serve as the power source, which enables electrochemical charge and discharge of the yarn to provide torsional rotation in opposite directions. In the simplest case, the researchers attach a paddle to the nanotube yarn, which enables torsional rotation to do useful work -- like mixing liquids on "micro-fluidic chips" used for chemical analysis and sensing.

The mechanism of torsional rotation is remarkable. Charging the nanotube yarns is like charging a supercapacitor -- ions migrate into the yarns to electrostatically balance the electronic charge electrically injected onto the nanotubes. Although the yarns are porous, this influx of ions causes the yarn to increase volume, shrink in length by up to a percent, and torsionally rotate. This surprising shrinkage in yarn length as its volume increases is explained by the yarn's helical structure, which is similar in structure to finger cuff toys that trap a child's fingers when elongated, but frees them when shortened.

Nature has used torsional rotation based on helically wound muscles for hundreds of millions of years, and exploits this action for such tasks as twisting the trunks of elephants and octopus limbs. In these natural appendages, helically wound muscle fibers cause rotation by contracting against an essentially incompressible, bone-less core. On the other hand, the helically wound carbon nanotubes in the nanotube yarns are undergoing little change in length, but are instead causing the volume of liquid electrolyte within the porous yarn to increase during electrochemical charging, so that torsional rotation occurs.

The combination of mechanical simplicity, giant torsional rotations, high rotation rates, and micron-size yarn diameters are attractive for applications, such as microfluidic pumps, valve drives, and mixers. In a fluidic mixer demonstrated by the researchers, a 15 micron diameter yarn rotated a 200 times larger radius and 80 times heavier paddle in flowing liquids at up to one rotation per second.

"The discovery, characterization, and understanding of these high performance torsional motors shows the power of international collaborations," said Ray H. Baughman, a corresponding author of the author of the Science article and Robert A. Welch Professor of Chemistry and director of The University of Texas at Dallas Alan G. MacDiarmid NanoTech Institute. "Researchers from four universities in three different continents that were born in eight different countries made critically important contributions."

Other co-authors of this article are Javad Foroughi (first author and research fellow), Geoffrey M. Spinks (a corresponding author and professor), and Gordon G. Wallace (professor) of the University of Wollongong in Australia; Jiyoung Oh (postdoctoral fellow), Mikhail E. Kozlov (research professor), and Shaoli Fang (research professor) at The University of Texas at Dallas; Tissaphern Mirfakhrai (postdoctoral fellow) and John D. W. Madden (professor) at The University of British Columbia; and Min Kyoon Shin (postdoctoral fellow) and Seon Jeong Kim (professor) at Hanyang University.

Funding for this research was provided by grants from the Air Force Office of Scientific Research, the Air Force AOARD program, the Office of Naval Research MURI program, and the Robert A. Welch Foundation in the United States; the Creative Research Initiative Center for Bio-Artificial Muscle in Korea; the Natural Sciences and Engineering Research Council of Canada; and the Australian Research Council.

'Robot Biologist' Solves Complex Problem from Scratch

One of the microformulators that the Wikswo lab has developed that will give ABE the ability to perform experiments without human intervention. (Credit: Courtesy of Wikswo Lab)

Science Daily  — First it was chess. Then it was Jeopardy. Now computers are at it again, but this time they are trying to automate the scientific process itself.

The paper that describes this accomplishment is published in the October issue of the journal Physical Biology and is currently available online.An interdisciplinary team of scientists at Vanderbilt University, Cornell University and CFD Research Corporation, Inc., has taken a major step toward this goal by demonstrating that a computer can analyze raw experimental data from a biological system and derive the basic mathematical equations that describe the way the system operates. According to the researchers, it is one of the most complex scientific modeling problems that a computer has solved completely from scratch.

The work was a collaboration between John P. Wikswo, the Gordon A. Cain University Professor at Vanderbilt, Michael Schmidt and Hod Lipson at the Creative Machines Lab at Cornell University and Jerry Jenkins and Ravishankar Vallabhajosyula at CFDRC in Huntsville, Ala.

The "brains" of the system, which Wikswo has christened the Automated Biology Explorer (ABE), is a unique piece of software called Eureqa developed at Cornell and released in 2009. Schmidt and Lipson originally created Eureqa to design robots without going through the normal trial and error stage that is both slow and expensive. After it succeeded, they realized it could also be applied to solving science problems.

One of Eureqa's initial achievements was identifying the basic laws of motion by analyzing the motion of a double pendulum. What took Sir Isaac Newton years to discover, Eureqa did in a few hours when running on a personal computer.

In 2006, Wikswo heard Lipson lecture about his research. "I had a 'eureka moment' of my own when I realized the system Hod had developed could be used to solve biological problems and even control them," Wikswo said. So he started talking to Lipson immediately after the lecture and they began a collaboration to adapt Eureqa to analyze biological problems.

"Biology is the area where the gap between theory and data is growing the most rapidly," said Lipson. "So it is the area in greatest need of automation."

Software passes test

The biological system that the researchers used to test ABE is glycolysis, the primary process that produces energy in a living cell. Specifically, they focused on the manner in which yeast cells control fluctuations in the chemical compounds produced by the process.

The researchers chose this specific system, called glycolytic oscillations, to perform a virtual test of the software because it is one of the most extensively studied biological control systems. Jenkins and Vallabhajosyula used one of the process' detailed mathematical models to generate a data set corresponding to the measurements a scientist would make under various conditions. To increase the realism of the test, the researchers salted the data with a 10 percent random error. When they fed the data into Eureqa, it derived a series of equations that were nearly identical to the known equations.

"What's really amazing is that it produced these equations a priori," said Vallabhajosyula. "The only thing the software knew in advance was addition, subtraction, multiplication and division."

Beyond Adam

The ability to generate mathematical equations from scratch is what sets ABE apart from Adam, the robot scientist developed by Ross King and his colleagues at the University of Wales at Aberystwyth. Adam runs yeast genetics experiments and made international headlines two years ago by making a novel scientific discovery without direct human input. King fed Adam with a model of yeast metabolism and a database of genes and proteins involved in metabolism in other species. He also linked the computer to a remote-controlled genetics laboratory. This allowed the computer to generate hypotheses, then design and conduct actual experiments to test them.

"It's a classic paper," Wikswo said.

In order to give ABE the ability to run experiments like Adam, Wikswo's group is currently developing "laboratory-on-a-chip" technology that can be controlled by Eureqa. This will allow ABE to design and perform a wide variety of basic biology experiments. Their initial effort is focused on developing a microfluidics device that can test cell metabolism.

"Generally, the way that scientists design experiments is to vary one factor at a time while keeping the other factors constant, but, in many cases, the most effective way to test a biological system may be to tweak a large number of different factors at the same time and see what happens. ABE will let us do that," Wikswo said.

The project was funded by grants from the National Science Foundation, National Institute on Drug Abuse, the Defense Threat Reduction Agency and the National Academies Keck Futures Initiative.

How the Zebra Gets Its Stripes: A Simple Genetic Circuit

 of University of California - San Diego)

Science Daily — Many living things have stripes, but the developmental processes that create these and other patterns are complex and difficult to untangle.

"The essential components can be buried in a complex physiological context," said Terence Hwa, a professor of physics at the University of California, San Diego, and one of the leaders of the study published October 14 in Science. "Natural systems make all kinds of wonderful patterns, but the problem is you never know what's really controlling it."Now a team of scientists has designed a simple genetic circuit that creates a striped pattern that they can control by tweaking a single gene.With genes taken from one species of bacterium and inserted into another, Hwa and colleagues from the University of Hong Kong assembled a genetic loop from two linked modules that senses how crowded a group of cells has become and responds by controlling their movements.

One of the modules secretes a chemical signal called acyl-homoserine lactone (AHL). As the bacterial colony grows, AHL floods the accumulating cells, causing them to tumble in place rather than swim. Stuck in the agar of their dish, they pile up.

Because AHL doesn't diffuse very far, a few cells escape and swim away to begin the process again.

Left to grow overnight, the cells create a target-like pattern of concentric rings of crowded and dispersed bacterial cells. By tweaking just one gene that limits how fast and far cells can swim, the researchers were able to control the number of rings the bacteria made. They can also manipulate the pattern by modifying how long AHL lasts before it degrades.

Although individual bacteria are single cells, as colonies they can act like a multicellular organism, sending and receiving signals to coordinate the growth and other functions of the colony. That means fundamental rules that govern the development of these patterns could well apply to critical steps in the development of other organisms.

To uncover these fundamental rules, Hwa and colleagues characterized the performance of their synthetic genetic circuit in two ways.

First, they precisely measured both the activity of individual genes in the circuit throughout the tumble-and-swim cycle. Then they derived a mathematical equation that describes the probability of cells flipping between swim and tumble motions.

Additional equations describe other aspects of the system, such as the dynamics of the synthesis, diffusion and deactivation of one of the cell-to-cell chemical signal AHL.

This three-pronged approach of "wet-lab" experiments, precise measurements of the results, and mathematical modeling of the system, characterize the emerging discipline of quantitative biology, Hwa said. "This is a prototype, a model of the kind of biology we want to do."

Co-authors include Jian-Dong Huang, associate professor of biochemistry at the University of Hong Kong additional researchers at Hong Kong Baptist University, the University of Marburg, and the University of Hong Kong including members of the 2008 iGEM team, which Hwa co-advised as a Distinguished Visiting Professor at UHK.

Hwa is a senior scientist with UC San Diego's Center for Theoretical Biological Physics.

Uncharted Territory: Scientists Sequence the First Carbohydrate Biopolymer

Structure of the bikunin: The portion on the left corresponds to the sugar part of the molecule, the sequence of which was determined in the current study. The portion on the right corresponds to the protein part of bikunin. (Credit: Rensselaer Polytechnic Institute)

ScienceDaily  — DNA and protein sequencing have forever transformed science, medicine, and society. Understanding the structure of these complex biomolecules has revolutionized drug development, medical diagnostics, forensic science, and our understanding of evolution and development. But, one major molecule in the biological triumvirate has remained largely uncharted: carbohydrate biopolymers.

The paper is titled "The proteoglycan bikunin has a defined sequence."

Today, for the first time ever, a team of researchers led by Robert Linhardt of Rensselaer Polytechnic Institute has announced in the October 9 Advanced Online Publication edition of the journal Nature Chemical Biology the sequence of a complete complex carbohydrate biopolymer. The surprising discovery provides the scientific and medical communities with an important and fundamental new view of these vital biomolecules, which play a role in everything from cell structure and development to disease pathology and blood clotting.

"Carbohydrate biopolymers, known as glycosaminoglycans, appear to be really important in how cells interact in higher organisms and could explain evolutionary differences and how development is driven. We also know that carbohydrate chains respond to disease, injury, and changes in the environment," said Linhardt, who is the Ann and John H. Broadbent Jr. '59 Senior Constellation Professor of Biocatalysis and Metabolic Engineering at Rensselaer. "In order to understand how and why this all happens, we first need to know their structure. And today, at least for the simplest glycosaminoglycan structure, we can now do this."

The first glycosaminoglycan sequenced was obtained from bikunin. Bikunin is a proteoglycan, a protein to which a single glycosaminoglycan chain is attached. Unlike less sophisticated carbohydrate biopolymers, such as starch and cellulose, the proteoglycans are decorated with structurally complex carbohydrates that enable them to perform more sophisticated and defined roles in the body. Bikunin, for example, is a natural anti-inflammatory that is used as a drug for the treatment of acute pancreatitis in Japan. It has the simplest chemical structure of any proteoglycan. Linhardt views the discovery of the structure of bikuin as the first step on the ladder to the discovery of the structure of more complex proteoglycans.

"The first genome sequences of DNA were on the simplest organisms such as bacteria. Once the technology was developed it ultimately led to the sequencing of the human genome," he said. "In our efforts to sequence carbohydrate biopolymers we don't yet know if the defined structure we observe for this simple protoglycan will hold for much more complex proteoglycans."

But, looking for structure in more complex proteoglycans will be among the next steps in the research for Linhardt and his team. The search for structure could help put to rest a long-running debate in the scientific community as to whether complex carbohydrate biopolymers require a defined structure to function.

"Despite all that is known about glycan formation, our understanding has not yet been deep enough to infer sequence or even determine if sequence occurs," Linhardt said. "These findings represent a new way of looking at these complex biomolecules as ordered structures."

Linhardt's research into carbohydrate sequencing began 30 years ago. In his previous work, he determined that some order existed in at least a portion of some carbohydrate biopolymers, but it did not represent the entire finished puzzle.

"Previously, we could see a pattern, but we could not see if all the chains were playing the same music. The tools did not yet exist. Now we can recognize it as a symphony."

To uncover the entire structure, Linhardt and his team, which was led by his doctoral student Mellisa Ly, borrowed a technique from the field of protein research called the proteomics top-down approach. As opposed to the bottom-up approach that first breaks apart a complex biopolymer into pieces and then rebuilds it piece by piece like a jigsaw puzzle, the top-down approach used by Linhardt and colleagues allows the researcher to picture the whole intact puzzle. This can only be accomplished with some of the most sophisticated technology available to the scientific community today, including very high-powered mass spectrometers.

Linhardt used a mass spectrometer located in the Rensselaer Center for Biotechnology and Interdisciplinary Studies (CBIS) to make his initial discoveries, and had these results independently confirmed on a separate and higher-level spectrometer at the University of Georgia. Mass spectrometers break down a molecule into separate charged particles or ions. These ions can then be categorized and analyzed based on their mass-to-charge ratio. These ratios then allow for sequencing of the entire molecule.

"This was truly the convergence of really sophisticated spectroscopy and its application to biology," Linhardt said. "We were fortunate to have a lot of time to play with the instrument at CBIS to understand its capabilities."

Beyond the technology it also took faith and determination. According to Linhardt, "It takes a student that is willing to try something even when the odds are pretty low. If it doesn't work, you make incremental progress. If it does work, you can make a great discovery. But, from the beginning you need to be a believer that it is worth taking the chance because it takes a lot of hard work in the lab."

And the odds weren't in Linhardt's favor. Despite being the most simple of proteoglycans, there were still 290 billion different possible sequences for the molecule.

"The first sample we looked at, we got the structure," Linhardt said. "In the end we did 15 chains and they all came back playing the same exact symphony."

The research is funded by the National Institutes of Health.

Linhardt and Ly were joined in the research by Tatiana Laremore of Rensselaer; Franklin Leach and Jonathan Amster of the University of Georgia; and Toshihiko Toida of Chiba University in Japan.

I’m Ready For More

“My dear Baladeva, best of our family, please come immediately with Your younger brother, Krishna. You both ate in the morning, and now You ought to eat something more.” (Mother Yashoda, Shrimad Bhagavatam, 10.11.16) No amount of affection previously offered can stop Mother Yashoda from continuing on with her motherly duties. It is not that once she performs a certain amount of work, she just sits back and relaxes the rest of the time. Rather, with each offering she makes to her children, her maternal affection only grows stronger. Though her beloved child, Lord Krishna, and His elder brother Balarama were properly fed in the morning, her worrying does not stop. To ensure that they continue to enjoy their youth properly, she agonizes over their well-being, thinking that they have not eaten enough. Therefore she calls them home, to return from the playground where the sacred pastimes of the Supreme Personality of Godhead take place. The young children ate their breakfast in the morning. From following the advice of Mother Yashoda, the queen of the farm community of Vraja, they had the strength to go out and play. Young children should be let free into the fields to enjoy their sportive tendencies. Rather than remain locked up, better the young bundles of energy be allowed to release their potential for activity. The fuel for their play comes from the mother’s love, which arrives in the form of her cooking. Mother Yashoda’s cooking is so tasty that even Krishna’s friends enjoy it. A young brahmana boy by the name of Madhumangala is known for coming over and eating more food than anyone else. Mother Yashoda loves this, as her hard work in the kitchen does not go to waste. She is ready to offer an endless amount of sumptuous delights to Krishna and His friends, and as a perfect match, Shri Krishna is willing to accept whatever His dear mother offers. Isn’t Lord Krishna considered the Supreme Personality of Godhead? If so, how can He have a mother and father? The more relevant question would be why shouldn’t He have parents if He so chooses? Why would that enjoyment be denied the Supreme Enjoyer? The Vedic tenets stipulate that God can be known by three primary characteristics. He is the original proprietor, the supreme enjoyer, and the best friend of the living entities. Enjoyment comes through association with His property and friends. In this area there can be different moods of association. Krishna is not picky; He will play the role perfectly to fit the particular devotional mood of His adherent. This even holds true with His enemies. How does this last part work? The enemies deny the existence of God. They come up with many justifications for their belief system. “Krishna is just a sectarian figure; evolution explains the creation; God is dead; religion is for those who can’t cope with death; God is made by man, not the other way around”, etc. Many excuses are made over the course of the many years of the earth’s existence. Krishna kindly reciprocates by remaining hidden from the vision of such people. If someone were to hate us and not want to see us, we would gladly oblige their request and stay away from their presence, especially if avoiding them would give us pleasure as well. Krishna is complete in Himself; therefore if someone is insistent on turning their back to Him, there is no loss on the Lord’s part. Krishna will find new ways to keep the miscreant further away from Him. “I am never manifest to the foolish and unintelligent. For them I am covered by My eternal creative potency [yoga-maya]; and so the deluded world knows Me not, who am unborn and infallible.” (Lord Krishna, Bhagavad-gita, 7.25) Why is such a person labeled a miscreant? Well, if God is the original proprietor responsible for providing the gifts we take for granted on a daily basis, wouldn’t anyone who tried to deny His existence be considered a miser? If I walk up to someone’s property and start pointing to different things and saying that they are mine, is that sane behavior? Yet this is precisely the tact followed by the staunch atheists, who continually observe material existence using their blunt sense perception and try to prove that God does not exist. They will always have the fuel necessary to continue in this endeavor, as Krishna will reveal bits and pieces of different aspects of His creation, with each successive discovery considered new and groundbreaking. Since the clock will always tick towards eventual death, the atheist is guaranteed to never acquire complete knowledge. Indeed, in the next life the same person gets to renew their search, with the slate wiped clean as far as knowledge goes. Just as Krishna engages with the atheist by remaining hidden, He fully appreciates the devotional efforts made by those willing to acknowledge the Lord’s existence. The more sincere the effort, the more information about God is revealed. As one ascends the chain of spiritual knowledge, they get to see more and more of Krishna. The most exalted devotees are those who get to interact with Krishna personally through a particular rasa, or transcendental mellow. In the beginning stages there is shanta-rasa, or venerable appreciation for the Supreme Lord. The fact that God is a person with spiritual attributes may not even be known in this stage, but at least He is respected. The concept of a “god-fearing” person correlates well with shanta-rasa. In shanta-rasa, the Supreme Lord’s blissful features are not well known. Not that these features can ever be absent, for Krishna is always inananda. He is always seen smiling and playing on His flute. As further devotional practice is followed, more of Krishna’s features are revealed, similar to how our vision clears up when more dust is removed from the eyes. The vision is what gets clearer, not the person being viewed. The steady engagement in bhakti-yoga, or devotional service, facilitated through chanting mantras like, “Hare Krishna Hare Krishna, Krishna Krishna, Hare Hare, Hare Rama Hare Rama, Rama Rama, Hare Hare”, helps one ascend to the higher platforms of spiritual interaction. Mother Yashoda and the residents of the farm community of Vrindavana are so elevated that they get to see Krishna directly. What’s even more remarkable is that Krishna uses His yoga-maya potency to keep His divinity hidden. Think of this as Krishna wanting to be treated just like one of the guys. He’s like everyone else after all, except His sparkling transcendental features cannot remain concealed. Therefore even when He appears on earth in a human form, His uniqueness is duly noted. In Vrindavana some five thousand years ago, Krishna crawled around on the floor like an ordinary infant. This behavior especially caught the interest of learned sages, who marveled at God’s ability to give the appearance of being an ordinary living entity. The purpose behind Krishna’s behavior was to grant the desire for interaction of the residents. Mother Yashoda particularly wanted interaction in vatsalya-rasa, or parental love. How amazing is Krishna? He is the Supreme Father, the fountainhead of all energies, the person from whom everything emanates, and yet He is kind enough to take on the role of a child to give pleasure to Nanda Maharaja and Mother Yashoda. For the interactions to be fully relished, Krishna behaved just like an ordinary boy, going out to the fields every day with His friends. Mother Yashoda was thus given the opportunity to feed her son, to smother Him in motherly affection. The accounts of Krishna’s pastimes are most thoroughly presented in the Shrimad Bhagavatam, which is also known as the Bhagavata Purana. Purana means “old”, so the Puranas are Vedic texts which expound upon the highest truths of life through stories relating to incidents from ancient times. Not that the events are fabricated, just the exact details provided are sequenced in such a way as to teach many lessons. The interactions between Mother Yashoda and her son show that God is always ready to accept more love from the devotees. Our responsibilities are not finished after muttering a few mantras and attending periodic religious functions. If we forget Krishna, we are automatically in a negative position. We don’t need the looming threat of punishment in hell to know that absence of God consciousness is detrimental. On the reverse side, remaining in Krishna’s company is always beneficial. Since God is absolute, His personal presence is not required. Just hearing about Mother Yashoda calling Krishna and Balarama to come home to eat is as good as witnessing the events firsthand. Prasadam cooking and distribution is modeled after this concept. The devotee follows the example of Mother Yashoda, knowing that the food they are offering to thedeity will be accepted by the Lord Himself, who never tires of eating anything offered with love and devotion. Mother Yashoda’s offerings fueled Krishna’s activities and they further bound her in a knot of loving affection to her son. In the same fashion, with every devotional act we take up, our attachment to Krishna tightens. Say Krishna’s name all the time and pretty soon you’ll see His beautiful, smiling face wherever you go. That image of the Lord holding His flute with His lotus-like hands and not showing the least stress on His face reminds the conditioned soul of what their constitutional position is. The Supreme Enjoyer is the resting place for those seeking enjoyment. The original proprietor kindly bestows gifts that can be used in furthering one’s God consciousness. The best friend of the living entities is waiting for His fragmental sparks to decide to come and play with Him. In the sacred land of Vrindavana, the food is provided by the cows and the grains, the preparations by Mother Yashoda, and the entertainment by Krishna and His sportive exploits. The ears are satisfied with the sounds emanating from the Lord’s flute and the eyes by His precious, adorable vision. With the complete package available to those who follow bhakti, why would anyone take to any other type of activity? The potential for becoming fully God conscious is the gift granted to the human being. Though the gift is there, one can’t enjoy it unless it is unwrapped. The young child doesn’t know how valuable the expensive vase in the house is; hence the parents warm them not to touch it. If not for this warning, the child may inadvertently throw the vase to the ground and break it. In a similar manner, the precious gift of the potential for tasting the fruit of one’s existence given to the human beings must be carefully unwrapped through the instructions of the spiritual master, or guru. The bona fide guru is a devotee of Krishna who always enjoys the Lord’s association in some way or another. Without the instruction of the guru, the precious gift will either remain unwrapped or get tossed aside as being unimportant. The guru will tell the student to regularly chant the holy names and hear the nectar of Krishna’s pastimes. Simply meditating on the one scene of Mother Yashoda calling Krishna and Balarama to come home can provide so much insight. From one tiny incident, so many transcendental thoughts can be awakened, leading the inquisitive mind towards the proper destination. The Shrimad Bhagavatam and its verses cannot be accurately priced; the information within is invaluable. Krishna is always ready to accept more love, so why shouldn’t we be the ones to offer it to Him? In Closing: “Krishna, I understand You want to go on playing, For I gave You plenty to eat this morning. But now it is time for You to come back home, You can’t survive on just the breakfast alone. Baladeva, it is time to come back with your brother, Do not you two hear the piteous cries of your mother?” In this way Mother Yashoda always looks to protect, Her son Krishna, who endless amount of love can accept. In these verses much wisdom and delight does abound, In the sacred Shrimad Bhagavatam of Vyasa they are found. Stay in Krishna consciousness, let not the mind drift, Tasting fruit of existence is mankind’s unique gift. To learn of this opportunity’s true value, Hear from guru, who speaks words of God that are true. Neglecting worship of Krishna do not make the mistake, In this life let the divine vision of Him the eyes take.