%0 Journal Article
%A Syson, Karl
%A Stevenson, Clare E. M.
%A Rashid, Abdul M.
%A Saalbach, Gerhard
%A Tang, Minhong
%A Tuukkanen, Anne
%A Svergun, Dmitri
%A Withers, Stephen G.
%A Lawson, David M.
%A Bornemann, Stephen
%T Structural Insight into How Streptomyces coelicolor Maltosyl Transferase GlgE Binds α-Maltose 1-Phosphate and Forms a Maltosyl-Enzyme Intermediate
%J Biochemistry
%V 53
%N 15
%@ 1520-4995
%C Columbus, Ohio
%I American Chemical Society
%M PUBDB-2015-01352
%P 2494 - 2504
%D 2014
%X GlgE (EC 2.4.99.16) is an α-maltose 1-phosphate:(1→4)-α- D -glucan 4-α- D -maltosyltransferase of theCAZy glycoside hydrolase 13_3 family. It is the definingenzyme of a bacterial α-glucan biosynthetic pathway and is agenetically validated anti-tuberculosis target. It catalyzes the α-retaining transfer of maltosyl units from α-maltose 1-phosphate to maltooligosaccharides and is predicted to use adouble-displacement mechanism. Evidence of this mechanismwas obtained using a combination of site-directed mutagenesis of Streptomyces coelicolor GlgE isoform I, substrate analogues,protein crystallography, and mass spectrometry. The X-ray structures of α-maltose 1-phosphate bound to a D394A mutein and aβ-2-deoxy-2-fluoromaltosyl-enzyme intermediate with a E423A mutein were determined. There are few examples of CAZyglycoside hydrolase family 13 members that have had their glycosyl-enzyme intermediate structures determined, and none beforenow have been obtained with a 2-deoxy-2-fluoro substrate analogue. The covalent modification of Asp394 was confirmed usingmass spectrometry. A similar modification of wild-type GlgE proteins from S. coelicolor and Mycobacterium tuberculosis was alsoobserved. Small-angle X-ray scattering of the M. tuberculosis enzyme revealed a homodimeric assembly similar to that of the S.coelicolor enzyme but with slightly differently oriented monomers. The deeper understanding of the structure−functionrelationships of S. coelicolor GlgE will aid the development of inhibitors of the M. tuberculosis enzyme.
%F PUB:(DE-HGF)16
%9 Journal Article
%U <Go to ISI:>//WOS:000334991100008
%$ pmid:24689960
%R 10.1021/bi500183c
%U https://bib-pubdb1.desy.de/record/207454