Protein Design and Protein Folding

Some of the tasks EGAD can perform are: prediction of mutation effects on protein stability and protein-complex formation to within ~1kcal/mol, automated scanning mutagenesis, including saturation mutagenesis, of proteins and protein complexes, total protein sequence design, design of ligand binding sites, optimization of sequences while considering multiple structures for the design of specific binding proteins and conformational switching, predicting the pK's of ionizable groups in proteins, and generating tables to display the distribution of energetic interactions in protein structures.

"The database aims to collect all folding data into one repository and, within the framework of the International Foldeomics Consortium (http://www.foldeomics.org), encourage sharing and data analysis. We are particularly interested in allowing the graphical analysis of raw-data on the site."

You can register to do advanced searches and to submit your own protein folding data.

While it is generally understood that proper protein folding happens spontaneously, scientists are still trying to understand what causes a particular protein to fold properly or go awry. The protein’s local environment plays a role, and other molecules in the neighborhood can assist or hinder the process, says Gierasch.

“It’s a delicate balance—proteins can tip to the dark side very readily,” she says.

Recently researchers have developed techniques that allow them to follow the fate of individual proteins within cells. Using an experimental set-up that they designed to do just that, Gierasch and Ignatova decided to see how the small molecule proline influenced protein folding. Proline is often taken up by cells in response to water loss, and other molecules with similar cellular roles have been shown to inhibit improper folding.

“There have been conflicting results,” says Gierasch, “In some cases these molecules inhibit misfolding, in some cases they promote it.”

This study was presented here on June 23rd at the Annual European Congress of Rheumatology (EULAR) on behalf of the Fibrillex Amyloid Secondary Trial (FAST) Group by Bouke P.C. Hazenberg, MD, coinvestigator and consultant in rheumatology, rheumatology, and clinical immunology at the University Medical Centre Groningen in Groningen, The Netherlands.

Amyloidosis is a protein-folding disorder that can occur in diseases characterized by extracellular deposition of insoluble fibrillar proteinaceous material, thereby resulting in loss of function of the tissue or organ involved.

In between the size of an atom and a drop of water, objects the size of living cells would be expected to vibrate mechanically at intermediate frequencies (10,000-500, 000 Hz).

But also consider that other oscillations besides mechanical shaking are of interest in cells, including the rate of protein folding and unfolding, the rate at which nutrients enter and leave a cell, the rate at which life processes take place, and many others.

Researchers at Northwestern University have offered a clue that may get to the core of the cell death question and establish a common mechanism in these diseases.

In a study, published in the journal Science, the research team showed that polyglutamine ( the toxic component of the protein responsible for Huntington's disease ) is so demanding on the cell's system that it changes the environment within the cell, causing other metastable, or partially folded, proteins to crash and lose function. Over time, this can cause the organism to die.

" Our results suggest that these disease-associated, aggregation-prone proteins may exert their destabilizing effects by interfering generally with other proteins that are having difficulty folding," said Richard I. Morimoto, who led the study.

Now, a discovery by Weill Cornell Medical College researchers is changing decades-old conventional wisdom on the "coiled coil," a common protein structural motif scientists thought they knew well.

The finding, recently published in the journal Structure, marks an incremental but important advance in understanding the complex, three-dimensional interactions between these building blocks of life.

"A deeper knowledge of these types of relationships is crucial, because it gets us a tiny step closer to recreating them artificially -- using 'designer proteins' to treat and even cure disease," explains senior author Dr. Min Lu, Associate Professor of Biochemistry at Weill Medical College of Cornell University, New York City.

While the sequencing of the human genome was justly hailed as a milestone, it may prove to be just a stop on the way to a much bigger prize -- protein engineering.

"Remember, genes are simply the code that cells use to make proteins," Dr. Lu explains.

Every day, the body creates tens of thousands of different proteins that fold together in complex, three-dimensional ways to perform specific functions.

"Understanding, fixing, and even reproducing these interrelationships is really the next scientific frontier, with infinite possibilities for the health sciences and beyond. Right now, however, we know very little about the way proteins fold," he says.

A series of discussions in a campus café in Lausanne has blossomed into an extraordinary collaboration between EPFL physics professor Paolo De Los Rios and University of Lausanne biology professor Pierre Goloubinoff. Using the principles of statistical physics, they have identified a simple, single mechanism that explains the mechanical role of molecular chaperones in protein folding and translocation, settling at the same time a long-standing controversy over this process.

Using plants, it is technically possible to grow, harvest, and process pharmaceutical proteins on a large scale. The genetically modified crop containing edible vaccine, for instance, can be consumed raw or partially processed. By targeting protein expression into specific organs of the plant such as grains, one creates a stable storage system. By targeting the recombinant protein to subcellular regions of the cells, such as the endoplasmic reticulum, a favorable environment is created for its appropriate folding and assembly, thus increasing the amount of recombinant proteins produced. Additionally, targeting the recombinant protein to the membrane concentrates the product. As a consequence, the cost of downstream purification is minimized. Finally, using plants for the production of drugs reduces the risk of contamination with human or animal pathogens, unlike production via animal or human sources.

So far, pharmaceutical proteins produced in plants can be classified into three groups: human biopharmaceutical proteins, including growth hormone, human serum albumin, β-interferon, and erythropoietin; recombinant antibodies such as IgG1 and IgM; and recombinant subunit vaccines, from the hepatitis B envelope proteins and the rabies virus glycoproteins to the cholera toxin B subunit.