4cbr
From Proteopedia
X-ray structure of the more stable human AGXT triple mutant (AGXT_HEM)
Structural highlights
DiseaseAGT1_HUMAN Primary hyperoxaluria type 1. The disease is caused by variants affecting the gene represented in this entry. FunctionAGT1_HUMAN Peroxisomal aminotransferase that catalyzes the transamination of glyoxylate to glycine and contributes to the glyoxylate detoxification (PubMed:10960483, PubMed:12777626, PubMed:24055001, PubMed:23229545, PubMed:26149463). Also catalyzes the transamination between L-serine and pyruvate and contributes to gluconeogenesis from the L-serine metabolism (PubMed:10347152).[1] [2] [3] [4] [5] [6] Publication Abstract from PubMedProtein stability is a fundamental issue in biomedical and biotechnological applications of proteins. Among them, gene- and enzyme-replacement strategies are promising approaches to treat inherited diseases that may benefit from protein engineering techniques, even though these beneficial effects have been largely unexplored. We apply here a sequence-alignment statistics procedure (consensus-based approach) to improve the activity and stability of the human alanine:glyoxylate aminotransferase (AGT) protein, an enzyme which causes primary hyperoxaluria type I (PH1) upon mutation. By combining only five consensus mutations, we obtain a variant (AGT-RHEAM) with largely enhanced in vitro thermal and kinetic stability, increased activity and no side effects on foldability and peroxisomal targeting in mammalian cells. The structure of AGT-RHEAM reveals changes at the dimer interface and improved electrostatic interactions responsible for increased kinetic stability. Consensus-based variants maintain the overall protein fold, crystallized more easily and improve the expression as soluble proteins in two different systems (AGT and CIPK24/SOS2). Thus, the consensus-based approach also emerges as a simple and generic strategy to increase the crystallization success for hard-to-get protein targets as well as to enhance protein stability and function for biomedical applications. The consensus-based approach for gene/enzyme replacement therapies and crystallization strategies: the case of human alanine:glyoxylate aminotransferase.,Mesa-Torres N, Yunta C, Fabelo-Rosa I, Gonzalez-Rubio JM, Sanchez-Ruiz JM, Salido E, Albert A, Pey AL Biochem J. 2014 Jun 24. PMID:24957194[7] From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine. See AlsoReferences
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