کد مقاله کد نشریه سال انتشار مقاله انگلیسی نسخه تمام متن
1177838 962625 2014 11 صفحه PDF دانلود رایگان
عنوان انگلیسی مقاله ISI
Structural characteristics of the nonallosteric human cytosolic malic enzyme
ترجمه فارسی عنوان
ویژگی های ساختاری آنزیم سیتوکسولیک منولی غیر آلسترتیک
کلمات کلیدی
آنزیم مالیک، فعال سازی آلوستریک، موتاژنز، سینتیک، سایت پیوند فومارات، پایداری ساختاری کواترنر
موضوعات مرتبط
مهندسی و علوم پایه شیمی شیمی آنالیزی یا شیمی تجزیه
چکیده انگلیسی


• The crystal structure of human c-NADP-ME in the apo form has been determined.
• The conformation of c-NADP-ME is similar to fumarate-bound m-NAD(P)-ME.
• The c-NADP-ME mutant (ME1_[51-105]) did not convert into an allosteric enzyme.
• Features of dimer interface highly correlate with the quaternary structural stability.

Human cytosolic NADP+-dependent malic enzyme (c-NADP-ME) is neither a cooperative nor an allosteric enzyme, whereas mitochondrial NAD(P)+-dependent malic enzyme (m-NAD(P)-ME) is allosterically activated by fumarate. This study examines the molecular basis for the different allosteric properties and quaternary structural stability of m-NAD(P)-ME and c-NADP-ME. Multiple residues corresponding to the fumarate-binding site were mutated in human c-NADP-ME to correspond to those found in human m-NAD(P)-ME. Additionally, the crystal structure of the apo (ligand-free) human c-NADP-ME conformation was determined. Kinetic studies indicated no significant difference between the wild-type and mutant enzymes in Km,NADP, Km,malate, and kcat. A chimeric enzyme, [51-105]_c-NADP-ME, was designed to include the putative fumarate-binding site of m-NAD(P)-ME at the dimer interface of c-NADP-ME; however, this chimera remained nonallosteric. In addition to fumarate activation, the quaternary structural stability of c-NADP-ME and m-NAD(P)-ME is quite different; c-NADP-ME is a stable tetramer, whereas m-NAD(P)-ME exists in equilibrium between a dimer and a tetramer. The quaternary structures for the S57K/N59E/E73K/S102D and S57K/N59E/E73K/S102D/H74K/D78P/D80E/D87G mutants of c-NADP-ME are tetrameric, whereas the K57S/E59N/K73E/D102S m-NAD(P)-ME quadruple mutant is primarily monomeric with some dimer formation. These results strongly suggest that the structural features near the fumarate-binding site and the dimer interface are highly related to the quaternary structural stability of c-NADP-ME and m-NAD(P)-ME. In this study, we attempt to delineate the structural features governing the fumarate-induced allosteric activation of malic enzyme.

ناشر
Database: Elsevier - ScienceDirect (ساینس دایرکت)
Journal: Biochimica et Biophysica Acta (BBA) - Proteins and Proteomics - Volume 1844, Issue 10, October 2014, Pages 1773–1783
نویسندگان
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