Scientists are discovering a binding secret behind the protein 'superglue & # 39;

Scientists from the Nanyang Technological University in Singapore (NTU Singapore), explained how a particular class of plant-derived enzymes, called peptide ligases, works to bind proteins. Such binding is an important process in drug development, for example by specifically binding a chemotherapy drug to an antibody that recognizes tumor markers to target cancer cells.

More generally, peptides ligases are a useful tool in biotechnological and biomedical applications such as labeling, imaging and protein monitoring in the body.

The scientists of the NTU Singapore have shown that the secret of the property of "superglue" a peptide ligase lies in two specific regions of the enzyme that gives it the ability to bind to others molecules and change the speed at which it acts.

The NTU team led by Associate Professor Julien Lescar and Professor James Tam of the NTU School of Biological Sciences used their new knowledge to develop a new peptide ligase created in the laboratory based on genetic information from Chinese violet (Viola yedoensis), a medicinal plant with antibiotic and anti-inflammatory properties.

The artificially created peptide ligase, also known as recombinant peptide ligase, can aid in the development of drugs made from components taken from living organisms because it overcomes the limitations of current methods, such as byproducts. or toxic molecules function and effectiveness of the drug.

The results were published in April in Proceedings of the National Academy of Sciences (PNAS).

Associate Professor Julien Lescar, also based at the NTU Institute for Structural Biology, said: "We used what we learned in nature to develop a recombinant enzyme in the laboratory It attaches itself to a specific protein, which then associates with another specific protein or molecule.This new approach could underpin better diagnostic tests or anti-cancer drugs. "

Currently, during the development of a drug, the protein molecules are chemically stitched. While this is effective, the process leaves by-products that can alter the function of the final product.

Plant-derived peptide ligases have been shown to be a more reliable protein than ligases derived from bacteria or the use of chemicals to bind proteins together. Professor Asscar, Lescar, noted that enzymes extracted from plants may contain traces of contaminants that could in turn trigger allergies, and that the availability of the enzyme depends on the success of the crop and harvest. In contrast, the new recombinant enzyme developed by NTU can be produced in large quantities in laboratories without any by-products.

Professor James Tam, also a member of NISB, said: "In very simple terms, our work in creating these ligases provides an improved platform for accurate bioproduction of drugs, diagnostics and biomaterials."

Take a leaf from the nature book

The NTU team studied the genetic information of enzymes extracted from Chinese violet (Viola yedoensis) and the violet of Canada (Viola canadensis). Instead of testing the efficiency of these extracted enzymes, scientists have developed five samples of recombinant enzymes by inserting the genes for these enzymes into an insect cell culture. Of the five samples, three are peptides ligases. The other two are proteases, which are enzymes that cleave protein molecules into smaller peptide chains.

Scientists have discovered that one of the recombinant peptide ligase samples, VyPAL2, has exceptional binding properties and that it is 3000 times more effective than three other types of known ligases.

Through a structural analysis of VyPAL2, the NTU team then reduced the "control centers" of its superglue property to two specific regions, called LAD1 and LAD2. LAD1 affects the level of enzymatic activity, while LAD2 determines whether the enzyme exhibits ligase or protease activity.

Transformation of proteases into peptides ligases

Another discovery derived from the knowledge of the molecular mechanism of peptide ligase is a method for transforming it from a cutter (a protease) into a linking agent (peptide ligase). This can be done by introducing mutations into the LAD1 and LAD2 regions of a protease.

Knowing this conversion process opens up possibilities for identifying interesting peptides-ligases by simple trawling in protein sequence databases, said Professor Asscar, Lescar.

"When you have tens of thousands of proteases, and only a few known ligase peptides, scanning the sequence databases with the LAD1 and LAD2 regions as a search criterion could lead to the discovery of more proteases that can be converted to ligase peptides." as a fishing expedition, but at least now we know where to fish. "

Future applications

The team recently received funding from NTUitive, NTU's innovation and enterprise company, and is currently working on developing the recombinant enzyme into one product. The product will eventually be sold to Epitoire, a start-up founded by Assoc Prof Lescar. The start-up sells DNA, RNA and protein reagents to academics and researchers wishing to modify their proteins.

The team also collaborates with local and foreign medical schools and health facilities to use this recombinant enzyme in diagnostic imaging, such as brain tumor imaging during a surgical procedure.

A patent has been filed for the creation of the recombinant enzyme, as well as for the mechanism that converts a protease into a ligase.