title: Thermus thermophilus glycoside hydrolase family 57 branching enzyme: crystal structure, mechanism of action, and products formed.
abstract: Branching enzyme (EC 2.4.1.18; glycogen branching enzyme; GBE) catalyzes the formation of α1,6-branching points in glycogen. Until recently it was believed that all GBEs belong to glycoside hydrolase family 13 (GH13). Here we describe the cloning and expression of the Thermus thermophilus family GH57-type GBE and report its biochemical properties and crystal structure at 1.35-Å resolution. The enzyme has a central (β/α)(7)-fold catalytic domain A with an inserted domain B between β2 and α5 and an α-helix-rich C-terminal domain, which is shown to be essential for substrate binding and catalysis. A maltotriose was modeled in the active site of the enzyme which suggests that there is insufficient space for simultaneously binding of donor and acceptor substrates, and that the donor substrate must be cleaved before acceptor substrate can bind. The biochemical assessment showed that the GH57 GBE possesses about 4% hydrolytic activity with amylose and in vitro forms a glucan product with a novel fine structure, demonstrating that the GH57 GBE is clearly different from the GH13 GBEs characterized to date.
authors.affiliation: Department of Microbiology, Groningen Biomolecular Sciences and Biotechnology Institute, Kerklaan 30, 9751 NN Haren, The Netherlands.
authors.forename: Marta
authors.initials: M
authors.lastname: Palomo
authors.forename: Tjaard
authors.initials: T
authors.lastname: Pijning
authors.forename: Thijs
authors.initials: T
authors.lastname: Booiman
authors.forename: Justyna M
authors.initials: JM
authors.lastname: Dobruchowska
authors.forename: Jeroen
authors.initials: J
authors.lastname: van der Vlist
authors.forename: Slavko
authors.initials: S
authors.lastname: Kralj
authors.forename: Antoni
authors.initials: A
authors.lastname: Planas
authors.forename: Katja
authors.initials: K
authors.lastname: Loos
authors.forename: Johannis P
authors.initials: JP
authors.lastname: Kamerling
authors.forename: Bauke W
authors.initials: BW
authors.lastname: Dijkstra
authors.forename: Marc J E C
authors.initials: MJ
authors.lastname: van der Maarel
authors.forename: Lubbert
authors.initials: L
authors.lastname: Dijkhuizen
authors.forename: Hans
authors.initials: H
authors.lastname: Leemhuis
mesh_terms: D015061:1,4-alpha-Glucan Branching Enzyme / Q000737:chemistry* / Q000378:metabolism; D020134:Catalytic Domain; D003001:Cloning, Molecular / Q000379:methods; D018360:Crystallography, X-Ray; D006026:Glycoside Hydrolases / Q000737:chemistry / Q000378:metabolism; D006868:Hydrolysis; D011487:Protein Conformation; D013379:Substrate Specificity; D016963:Thermus thermophilus / Q000201:enzymology*
chemical_list: D015061:1,4-alpha-Glucan Branching Enzyme; D006026:Glycoside Hydrolases