Carrasco Patricia, Menao Sebastian, López-Viñas Eduardo, Santpere Gabriel, Clotet Josep, Sierra Adriana Y, Gratacós Esther, Puisac Beatriz, Gómez-Puertas Paulino, Hegardt Fausto G, Pie Juan, Casals Núria
Department of Biochemistry and Molecular Biology, School of Health Sciences, Universitat Internacional de Catalunya, E-08195 Sant Cugat, Barcelona, Spain.
Mol Genet Metab. 2007 Jun;91(2):120-7. doi: 10.1016/j.ymgme.2007.03.007. Epub 2007 Apr 24.
3-hydroxy-3-methylglutaryl-CoA (HMG-CoA) lyase adopts a (betaalpha)(8) TIM barrel structure with an additional beta9, alpha11 and alpha12 helices. Location of HMG part of the substrate has been suggested but the binding mode for the CoA moiety remains to be resolved. As mutation F305 fs(-2), which involves the last 21 residues of the protein, and mutation K48N caused 3-hydroxy-3-methylglutaric aciduria in two patients, we examined the role of the C-terminal end and Lys(48) in enzyme activity. Expression studies of various C-terminal-end-deleted and K48N-mutated proteins revealed that residues 311-313 (localized in the loop between alpha11 and alpha12 helices) and Lys(48) are essential for enzyme activity. An in silico docking model locating HMG-CoA on the surface of the enzyme implicates Asn(311) and Lys(313) in substrate binding by establishing multiple polar contacts with phosphate and ribose groups of adenosine, and Lys(48) by contacting the carboxyl group of the panthotenic acid moiety.
3-羟基-3-甲基戊二酰辅酶A（HMG-CoA）裂解酶采用（βα）8 桶状结构，还有额外的β9、α11和α12螺旋。底物的HMG部分的定位已有人提出，但辅酶A部分的结合模式仍有待确定。由于涉及蛋白质最后21个残基的F305 fs（-2）突变以及K48N突变在两名患者中导致了3-羟基-3-甲基戊二酸尿症，我们研究了C末端和赖氨酸48在酶活性中的作用。对各种C末端缺失和K48N突变蛋白的表达研究表明，残基311 - 313（位于α11和α12螺旋之间的环中）和赖氨酸48对酶活性至关重要。通过计算机对接模型将HMG-CoA定位在酶表面，表明天冬酰胺311和赖氨酸313通过与腺苷的磷酸和核糖基团建立多个极性接触参与底物结合，而赖氨酸48则通过与泛酸部分的羧基接触参与底物结合。
