Posts tagged proteins
Articles and news from the latest research reports.
Using bioluminescent proteins from a jellyfish, a team of scientists has lit up the inside of a neuron, capturing spectacular video footage that shows the movement of proteins throughout the cell.
(Source: medicalxpress.com)
Next Generation: In Vivo Drug Factories
By Sabrina Richards | August 13, 2012
Researchers use UV light to stimulate protein production in nano-sized delivery capsules in mice.
Nanoparticles expressing a GFP reporter.
Device: Science is one step closer to producing drugs in the right place at the right time in the body, avoiding the collateral damage of untargeted treatments. Researchers led by Daniel Anderson at the Massachusetts Institute of Technology have designed nanoparticles that can be stimulated via UV light to produce proteins on demand in vivo.
The new method, which involves packaging the molecular machinery for making proteins into a membraned capsule, allows the researchers to spatially and temporally regulate protein production, said Zhen Gu, who also researches nanoparticle drug delivery at North Carolina State and University of North Carolina, Chapel Hill, but did not participate in the research. “They can control generation of a protein at any time with a trigger of light.”
The scientists created the nano-sized “protein factories” by using lipids to encapsulate polymerase and other machinery necessary for protein production from E. coli, along with a DNA plasmid containing a gene of interest. To block transcription until the right moment, they added a DNA “photo-labile cage” to the plasmid—a small chemical that inhibits transcription but is cleaved by exposure to UV light.
To test the principle in vivo, the researchers used luciferase as the reporter protein and injected mice with the nanovesicles. After zapping them with UV light at the site of injection, they were able to measure a local burst of luminescence.
What’s new: Protein expression in liposomes has been possible for at least 10 years, said Mitchel Doktycz, a synthetic biologist at Oak Ridge National Laboratory in Tennessee. What is new, said Doktycz, who did not participate in the research, is being able to control the timing of protein expression in an animal. “They can do it remotely,” he said.
And that switch is not limited to UV light, added Gu, but will likely work with other wavelengths using different chemical ligands.
Avi Schroeder
Importance: Many life-saving drugs, such as chemotherapy, can have nasty and toxic effects outside the tissues they’re designed to treat. The goal of remotely-controlled factories like Anderson’s is to produce a drug in a specific place (such as a tumor) at a specific time (after enough particles have accumulated to produce a therapeutic effect). Anderson’s group is “trying to deliver a payload, [and] activate [it] in a specific spot, so they’re not dosing everywhere,” Doktycz explained—which has the potential to minimize side effects while maximizing therapeutic benefit.
Needs improvement: “We have a long way to go still before we have a drug factory that will land in a target tissue to produce a drug of interest,” noted Anderson. The study has proved the principle of the first step—getting the protein expressed on signal—but future research will need to ensure that the nanoparticles and the proteins they produce aren’t toxic in the wrong place, and that they get to the right location. Targeting the nanoparticles to the appropriate tissues might be achieved by “decorating” the surface of the vesicles with specific proteins, said Gu.
Furthermore, although most of the materials in the current particles are probably safe, some are microorganism-derived, Anderson pointed out, and most likely need to be switched to human alternatives. Finally, getting the drug expressed is also just one part of the problem, said Doktycz. So far the system has no way to re-cage the DNA to halt protein production when it’s no longer needed. “Turning on is one thing, but turning off is another,” he said.
(Source: the-scientist.com)
The genetic code of the fruit fly Drosophila has been hacked into, allowing it to make proteins with properties that don’t exist in the natural world. The advance could ultimately lead to the creation of new or “improved” life forms in the burgeoning field of synthetic biology.
The four letters of the genetic code, A, C, T and G, are read in triplets, called codons, by the cell’s protein-making machinery. Each codon gives an instruction for the type of amino acid that gets added next in a protein chain, or tells the machinery to stop.
Complex proposition
As a proof of principle, Chin’s team has engineered fruit flies that incorporated three new amino acids into proteins in the cells of their ovaries.
The flies were engineered using bacteria that had been modified to insert the genetic code for the unnatural amino acid into the fly DNA. There was no apparent impact on the flies’ health, and they even produced healthy offspring that also made the new protein chains.
Bulletproof flies
None of the amino acids were particularly remarkable, but the fact that engineering the flies had no obvious impact on their health suggests that many more useful amino acids could be similarly incorporated.
For example, work in bacterial cells has shown that it is possible to incorporate unnatural amino acids that cross-link to each other or turn an enzyme’s activity on or off when a light is shone on them. Doing this in a complex organism like a fly could shed new light on how proteins interact within cells, or how rapidly turning an enzyme on or off affects the cell’s function.
The technique could even be used to create animals with new or improved properties, although that is probably some years off.
Natural birth — but not C-section — triggers brain boosting proteins
Vaginal birth triggers the expression of a protein in the brains of newborns that improves brain development and function in adulthood, according to a new study by Yale School of Medicine researchers, who also found that this protein expression is impaired in the brains of offspring delivered by caesarean section (C-sections).
These findings are published in the August issue of PLoS ONE by a team of researchers led by Tamas Horvath, the Jean and David W. Wallace Professor of Biomedical Research and chair of the Department of Comparative Medicine at Yale School of Medicine.
The team studied the effect of natural and surgical deliveries on mitochondrial uncoupling protein 2 (UCP2) in mice. UCP2 is important for the proper development of hippocampal neurons and circuits. This area of the brain is responsible for short- and long-term memory. UCP2 is involved in cellular metabolism of fat, which is a key component of breast milk, suggesting that induction of UCP2 by natural birth may aid the transition to breast feeding.
The researchers found that natural birth triggered UCP2 expression in the neurons located in the hippocampal region of the brain. This was diminished in the brains of mice born via C-section. Knocking out the UCP2 gene or chemically inhibiting UCP2 function interfered with the differentiation of hippocampal neurons and circuits, and impaired adult behaviors related to hippocampal functions.
“These results reveal a potentially critical role of UCP2 in the proper development of brain circuits and related behaviors,” said Horvath. “The increasing prevalence of C-sections driven by convenience rather than medical necessity may have a previously unsuspected lasting effect on brain development and function in humans as well.”
Fragile X and Down Syndromes Share Signalling Pathway for Intellectual Disability
"We have shown for the first time that some of the proteins altered in Fragile X and Down syndromes are common molecular triggers of intellectual disability in both disorders," said Kyung-Tai Min, one of the lead authors of the study and a professor at Indiana University and the Ulsan National Institute of Science and Technology in Korea. "Specifically, two proteins interact with each other in a way that limits the formation of spines or protrusions on the surface of dendrites." He added: "These outgrowths of the cell are essential for the formation of new contacts with other nerve cells and for the successful transmission of nerve signals. When the spines are impaired, information transfer is impeded and mental retardation takes hold."
To investigate membrane fusion during synaptic transmission (top), Rothman, Pincet, and colleagues designed an artificial version of the event. They exposed lipid nanodiscs embedded with SNARE proteins to vesicles containing complementary SNARE proteins. Only one SNARE protein complex was required for fusion between the discs and vesicles (A), but three were necessary to create a stable pore to release the neurotransmitter contained within the vesicle (B).
How a protein meal tells your brain you are full
July 5, 2012
Feeling full involves more than just the uncomfortable sensation that your waistband is getting tight. Investigators reporting online on July 5th in the Cell Press journal Cell have now mapped out the signals that travel between your gut and your brain to generate the feeling of satiety after eating a protein-rich meal. Understanding this back and forth loop between the brain and gut may pave the way for future approaches in the treatment and/or prevention of obesity.
Feeling full involves more than just the uncomfortable sensation that your waistband is getting tight. Investigators reporting online on July 5th in the Cell Press journal Cell have now mapped out the signals that travel between your gut and your brain to generate the feeling of satiety after eating a protein-rich meal. Understanding this back and forth loop between the brain and gut may pave the way for future approaches in the treatment and/or prevention of obesity. Credit: Duraffourd et al., Cell
Food intake can be modulated through mu-opioid receptors (MORs, which also bind morphine) on nerves found in the walls of the portal vein, the major blood vessel that drains blood from the gut. Specifically, stimulating the receptors enhances food intake, while blocking them suppresses intake. Investigators have now found that peptides, the products of digested dietary proteins, block MORs, curbing appetite. The peptides send signals to the brain that are then transmitted back to the gut to stimulate the intestine to release glucose, suppressing the desire to eat.
Mice that were genetically engineered to lack MORs did not carry out this release of glucose, nor did they show signs of ‘feeling full’, after eating high-protein foods. Giving them MOR stimulators or inhibitors did not affect their food intake, unlike normal mice.
Because MORs are also present in the neurons lining the walls of the portal vein in humans, the mechanisms uncovered here may also take place in people.
"These findings explain the satiety effect of dietary protein, which is a long-known but unexplained phenomenon,” says senior author Dr. Gilles Mithieux of the Université de Lyon, in France. “They provide a novel understanding of the control of food intake and of hunger sensations, which may offer novel approaches to treat obesity in the future,” he adds.
Provided by Cell Press
Source: medicalxpress.com
Researchers identify new group of proteins in the brains of Alzheimer’s patients
June 13, 2012
Researchers from Boston University School of Medicine (BUSM) have identified a novel group of proteins that accumulate in the brains of patients with Alzheimer’s disease. These findings, which appear online in the Journal of Neuroscience, may open up novel approaches to diagnose and stage the progression likelihood of the disease in Alzheimer patients.
Alzheimer’s disease is presumed to be caused by the accumulation of β-amyloid, which then induces aggregation of a neuronal protein, called tau, and neurodegeneration ensues. The diagnosis of Alzheimer’s disease focuses on β-amyloid and tau protein, with much attention focusing on radiolabeled markers that bind to β-amyloid (such as the compound PiB). However, imaging β-amyloid is problematic because many cognitively normal elderly have large amounts of β-amyloid in their brain, and appear as “positives” in the imaging tests.
Therapeutic approaches for Alzheimer’s disease generally have focused on β-amyloid because the process of producing a neurofibrillary tangle composed on tau protein has been poorly understood. Hence, few tau therapies have been developed. According to the researchers, this study makes important advances on both of these fronts.
The BUSM researchers identified a new group of proteins, termed RNA-binding proteins, which accumulate in the brains of patients with Alzheimer’s disease, and are present at much lower levels in subjects who are cognitively intact. The group found two different proteins, both of which show striking patterns of accumulation. “Proteins such as TIA-1 and TTP, accumulate in neurons that accumulate tau protein, and co-localize with neurofibrillary tangles. These proteins also bind to tau, and so might participate in the disease process,” explained senior author Benjamin Wolozin, MD, PhD, a professor in the departments of pharmacology and neurology at BUSM. “A different RNA binding protein, G3BP, accumulates primarily in neurons that do not accumulate pathological tau protein. This observation is striking because it shows that neurons lacking tau aggregates (and neurofibrillary tangles) are also affected by the disease process,” he added.
The researchers believe this work opens up novel approaches to diagnose and stage the likelihood of progression by quantifying the levels of these RNA-binding protein biomarkers that accumulate in the brains of Alzheimer patients.
Wolozin’s group also pursued the observation that some of the RNA binding proteins bind to tau protein, and tested whether one of these proteins, TIA-1, might contribute to the disease process. Previously, scientists have demonstrated that TIA-1 spontaneously aggregates in response to stress as a normal part of the stress response. Wolozin and his colleagues hypothesize that since TIA-1 binds tau, it might stimulate tau aggregation during the stress response. They introduced TIA-1 into neurons with tau protein, and subjected the neurons to stress. Consistent with their hypothesis, tau spontaneously aggregated in the presence of TIA-1, but not in the absence. Thus, the group has potentially identified an entirely novel mechanism to induce tau aggregates de novo. In future work, the group hopes to use this novel finding to understand how neurofibrillary tangles for in Alzheimer’s disease and to screen for novel compounds that might inhibit the progression of Alzheimer’s disease.
Provided by Boston University Medical Center
Source: medicalxpress.com
New Molecules Important for Vision and Brain Function Identified
ScienceDaily (June 11, 2012) — In a pair of related studies, scientists from the Florida campus of The Scripps Research Institute have identified several proteins that help regulate cells’ response to light — and the development of night blindness, a rare disease that abolishes the ability to see in dim light.
In the new studies, published recently in the journals Proceedings of the National Academy of Sciences (PNAS) and The Journal of Cell Biology, Scripps Florida scientists were able to show that a family of proteins known as Regulator of G protein Signaling (RGS) proteins plays an essential role in vision in a dim-light environment.
"We were looking at the fundamental mechanisms that shape our light sensation," said Kirill Martemyanov, a Scripps Research associate professor who led the studies. "In the process, we discovered a pair of molecules that are indispensible for our vision and possibly play critical roles in the brain."
In the PNAS study, Martemyanov and his colleagues identified a pair of regulator proteins known as RGS7 and RGS11 that are present specifically in the main relay neurons of the retina called the ON-bipolar cells. “The ON-bipolar cells provide an essential link between the retinal light detectors — photoreceptors and the neurons that send visual information to the brain,” explained Martemyanov. “Stimulation with light excites these neurons by opening the channel that is normally kept shut by the G proteins in the dark. RGS7 and RGS11 facilitate the G protein inactivation, thus promoting the opening of the channel and allowing the ON-bipolar cells to transmit the light signal. It really takes a combined effort of two RGS proteins to help the light overcome the barrier for propagating the excitation that makes our dim vision possible.”
In the Journal of Cell Biology study, Martemyanov and his colleagues unraveled another key aspect of the RGS7/RGS11 regulatory response — they identified a previously unknown pair of orphan G protein-coupled receptors (GPCRs) that interact with these RGS proteins and dictate their biological function.
GPCRs are a large family of more than 700 proteins, which sit in the cell membrane and sense various molecules outside the cell, including odors, hormones, neurotransmitters, and light. After binding these molecules, GPCRs trigger the appropriate response inside the cell. However, for many GPCRs the activating molecules have not yet been identified and these are called “orphan” receptors.
The Martemyanov group has found that two orphan GPCRs — GPR158 and GPR179 — recruit RGS proteins and thus help serve as brakes for the conventional GPCR signaling rather than play an active signaling role.
In the case of retinal ON-bipolar cells, GPR179 is required for the correct localization of RGS7 and RGS11. Their mistargeting in animal models lacking GPR179 or human patients with mutations in the GPR179 gene may account for their night blindness, according to the new study. Intriguingly, in the brain GPR158 appears to play a similar role in localizing RGS proteins, but instead of contributing to vision, it helps RGS proteins regulate the m-opioid receptor, a GPCRs that mediates pleasurable and pain-killing effects of opioids.
"We are really in the very beginning of unraveling this new biology and understanding the role of discovered orphan GPR158/179 in regulation of neurotransmitter signaling in the brain and retina," Martemyanov said. "The hope is that better understanding of these new molecules will lead to the design of better treatments for addictive disorders, pain, and blindness."
Source: Science Daily
Speeding Up Drug Discovery With Rapid 3-D Mapping of Proteins
ScienceDaily (May 30, 2012) — A new method for rapidly solving the three-dimensional structures of a special group of proteins, known as integral membrane proteins, may speed drug discovery by providing scientists with precise targets for new therapies, according to a paper published May 20 in Nature Methods.
Using their new rapid technique, Choe’s team generated the structure of a hIMP known as TMEM14A, shown here in multiple three-dimensional conformations. (Credit: Courtesy of the Salk Institute for Biological Studies)
The technique, developed by scientists at the Salk Institute for Biological Studies, provides a shortcut for determining the structure of human integral membrane proteins (hIMPs), molecules found on the surface of cells that serve as the targets for about half of all current drugs.
Knowing the exact three-dimensional shape of hIMPs allows drug developers to understand the precise biochemical mechanisms by which current drugs work and to develop new drugs that target the proteins.
"Our cells contain around 8,000 of these proteins, but structural biologists have known the three-dimensional structure of only 30 hIMPs reported by the entire field over many years," says Senyon Choe, a professor in Salk’s Structural Biology Laboratory and lead author on the paper. "We solved six more in a matter of months using this new technique. The very limited information on the shape of human membrane proteins hampers structure-driven drug design, but our method should help address this by dramatically increasing the library of known hIMP structures."
Integral membrane proteins are attached to the membrane surrounding each cell, serving as gateways for absorbing nutrients, hormones and drugs, removing waste products, and allowing cells to communicate with their environment. Many diseases, including Alzheimer’s, heart disease and cancer have been linked to malfunctioning hIMPs, and many drugs, ranging from aspirin to schizophrenia medications, target these proteins.
Most of the existing drugs were discovered through brute force methods that required screening thousands of potential molecules in laboratory studies to determine if they had a therapeutic effect. Given a blueprint of the 3D structure of a hIMP involved in a specific disease, however, drug developers could focus only on molecules that are most likely to interact with the target hIMP, saving time and expense.
In the past, it was extremely difficult to solve the structure of hIMPs, due to the difficulty of harvesting them from cells and the difficulty of labeling the amino acids that compose the proteins, a key step in determining their three-dimensional configuration.
"One problem was that hIMPs serve many functions in a cell, so if you tried to engineer cells with many copies of the proteins on their membrane, they would die before you could harvest the hIMPs," says Christian Klammt, a postdoctoral researcher in Choe’s lab and a first author on the paper.
To get around this, the scientists created an outside-the-cell environment, called cell-free expression system, to synthesize the proteins. They used a plexiglass chamber that contained all the biochemical elements necessary to manufacture hIMPs as if they were inside the cell. This system provided the researchers with enough of the proteins to conduct structural analysis.
The cell-free method also allowed them to easily add labeled amino acids into the biochemical stew, which were then incorporated into the proteins. These amino acids gave off telltale structural clues when analyzed with nuclear magnetic resonance spectroscopy, a method for using the magnetic properties of atoms to determine a molecule’s physical and chemical properties.
"It was very difficult and inefficient to introduce labeled amino acids selectively into the protein produced in live cells," says Innokentiy Maslennikov, a Salk staff scientist and co-first author on the paper. "With a cell-free system, we can precisely control what amino acids are available for protein production, giving us isotope-labeled hIMPs in large quantities. Using a proprietary labeling strategy we devised a means to minimize the number of samples to prepare."
Prior methods might take up to a year to determine a single protein structure, but using their new method, the Salk scientists determined the structure of six hIMPs within just 18 months. They have already identified 38 more hIMPs that are suitable for analysis with their technique, and expect it will be used to solve the structure for many more.
Source: Science Daily