Dual Coenzyme Specificity of Photosynthetic Glyceraldehyde-3-phosphate Dehydrogenase Interpreted by the Crystal Structure of A4 Isoform Complexed with NAD

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• 作者：Giuseppe Falini ; Simona Fermani ; Alberto Ripamonti ; Piera Sabatino ; Francesca Sparla ; Paolo Pupillo ; and Paolo Trost
• 刊名：Biochemistry
• 出版年：2003
• 出版时间：April 29, 2003
• 年：2003
• 卷：42
• 期：16
• 页码：4631 - 4639
• 全文大小：488K
• 年卷期：v.42,no.16(April 29, 2003)
• ISSN：1520-4995

文摘

Photosynthetic glyceraldehyde-3-phosphate dehydrogenase (GAPDH) of Spinacia oleraceabelongs to a wide group of GAPDHs found in most organisms displaying oxygenic photosynthesis, includingcyanobacteria, green and red algae, and higher plants. As a major catalytic difference with respect toglycolytic GAPDH, photosynthetic GAPDH exhibits dual cofactor specificity toward pyridine nucleotideswith a preference for NADP(H). Here we report the crystal structure of NAD-complexed recombinantA₄-GAPDH (NAD-A₄-GAPDH) from Spinacia oleracea, expressed in Escherichia coli. Its superimpositiononto native A₄-GAPDH complexed with NADP (NADP-A₄-GAPDH) pinpoints specific conformationalchanges resulting from cofactor replacement. In photosynthetic NAD-A₄-GAPDH, the side chain of Asp32is oriented toward the coenzyme to interact with the adenine ribose diol, similar to glycolytic GAPDHs(NAD-specific). On the contrary, in NADP-A₄-GAPDH Asp32 moves away to accommodate the additional2'-phosphate group of the coenzyme and to minimize electrostatic repulsion. Asp32 rotation is allowedby the presence of the small residue Ala40, conserved in most photosynthetic GAPDHs, replacing bulkyamino acid side chains in glycolytic GAPDHs. While in NADP-A₄-GAPDH two amino acids, Thr33 andSer188, are involved in hydrogen bonds with the 2'-phosphate group of NADP, in the NAD-complexedenzyme these interactions are lacking. The crystallographic structure of NAD-A₄-GAPDH highlights thatfour residues, Thr33, Ala40, Ser188, and Ala187 (Leu, Leu, Pro, and Leu respectively, in glycolyticBacillus stearothermophilus GAPDH sequence) are of primary importance for the dual cofactor specificityof photosynthetic GAPDH. These modifications seem to trace the minimum evolutionary route for aprimitive NAD-specific GAPDH to be converted into the NADP-preferring enzyme of oxygenicphotosynthetic organisms.