BioChem - DOS

Structural insights into the substrate specificity and activity of a novel mannose 2-epimerase from Runella slithyformis

Acta crystallographica. Section D, Structural biology

Wang, Hang; Sun, Xiaomei; Saburi, Wataru; Hashiguchi, Saki; Yu, Jian; Ose, Toyoyuki; Mori, Haruhide; Yao, Min

<P>Mannose 2-epimerase (ME), a member of the acylglucosamine 2-epimerase (AGE) superfamily that catalyzes epimerization of D-mannose and D-glucose, has recently been characterized to have potential for D-mannose production. However, the substrate-recognition and catalytic mechanism of ME remains unknown. In this study, structures of <I>Runella slithyformis</I> ME (<I>Rs</I>ME) and its D254A mutant [<I>Rs</I>ME(D254A)] were determined in their apo forms and as intermediate-analog complexes [<I>Rs</I>ME-D-glucitol and <I>Rs</I>ME(D254A)-D-glucitol]. <I>Rs</I>ME possesses the (&alpha;/&alpha;)6-barrel of the AGE superfamily members but has a unique pocket-covering long loop (loop&alpha;7-&alpha;8). The <I>Rs</I>ME-D-glucitol structure showed that loop&alpha;7-&alpha;8 moves towards D-glucitol and closes the active pocket. Trp251 and Asp254 in loop&alpha;7-&alpha;8 are only conserved in MEs and interact with D-glucitol. Kinetic analyses of the mutants confirmed the importance of these residues for <I>Rs</I>ME activity. Moreover, the structures of <I>Rs</I>ME(D254A) and <I>Rs</I>ME(D254A)-D-glucitol revealed that Asp254 is vital for binding the ligand in a correct conformation and for active-pocket closure. Docking calculations and structural comparison with other 2-epimerases show that the longer loop&alpha;7-&alpha;8 in <I>Rs</I>ME causes steric hindrance upon binding to disaccharides. A detailed substrate-recognition and catalytic mechanism for monosaccharide-specific epimerization in <I>Rs</I>ME has been proposed.</P>

2023

International Union of Crystallography

0907-4449

2059-7983

79

7

pp.585-595

10.1107/s205979832300390x

http://click.ndsl.kr/servlet/OpenAPIDetailView?keyValue=05787966&target=NART&cn=NART125218256