Community Research and Development Information Service - CORDIS

Final Report Summary - ASR-COMPENZDES (Active Site Repurposing – computational design of newenzyme functionalities by emulating nature)

Proteins, nature's building blocks of live, have distinct three-dimensional structures that are made out of a string of amino acids. In nature there are twenty different types of amino acids that are used to make a protein. In a highly regulated cellular process the amino acids are strung together and arranged into a distinct structure to make proteins. The amino acid sequence is also called the proteins primary structure. It tends to build stable local structure elements, called secondary structure. Those can be spiral structures (helices), elongated stretches (strands) or swirly random structures (loops). With only these twenty building blocks and three structure motifs, nature can build all the proteins we know, through a processes called protein folding, in which the string of amino acids folds into helices, strands and loops, which in turn fold down onto each other to form a stable configuration that represents a low energy state for a particular amino acid sequence. Remarkably, even for small protein structures, this process would take the present time of the universe, if all the conformations it could adopt were explored (commonly referred to as Levinthal's paradox or the protein folding problem). This also means, if we can solve the protein folding problem efficiently, we could make proteins from scratch according to our needs.
During the last decades, proteins found wide spread use as drugs to cure diseases or to generate highly pure chemical compounds as well as to degrade environmental pollutants, while at the same time they produce only minimal amounts of waste and exhibit an excellent resource balance. However, for present day applications of protein based nano-materials and general protein engineering applications, very stable building blocks are desirable, but naturally occurring proteins are only marginally stable. We developed a general procedure for designing new protein structures by taking a set of equations first derived by Francis Crick in 1953, which accurately describe helical structures and combining them with sophisticated computational modeling tools. This enabled us to generate new helical protein structures of more or less arbitrary size and with unprecedented stabilities. Our designs are stable above 95°C - a temperature at which most natural proteins have long started to degrade - and in other highly degrading conditions. Only the combination of high temperatures around 100°C and chemicals that degrade proteins made the designs fall apart. This makes our proteins some of the most stable ones ever to be described to date. We want to emphasize that this is an excellent example of how relatively old discoveries can be combined with modern day techniques to build marvelous new things. We are now using this computational approach to custom design hyperstable helical proteins with fine-tuned geometries for a range of biomedical and biotechnological applications. Most interesting to us is their catalytic functionalization, which we are currently working on. This functionalization is facilitated by an approach that we call de novo active-site repurposing. Here, we analyzed naturally occurring binding sites, which are capable of catalyzing a similar reaction to the one we seek and repurpose this machinery to bind a new substrate and/or catalyze another reaction. We have shown that this is possible starting from natural protein structures belonging to the same enzyme-family, with the particular example of an aromatic, nucleophilic substitution reaction that catalyzes the breakdown of a herbicidal compound.

Reported by

UNIVERSITAET GRAZ
Austria

Subjects

Life Sciences
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