Characterization and crystal structure determination of β‐1,2‐mannobiose phosphorylase from Listeria innocua

Abstract

Edited by Judit OvádiKeywords: b-1,2-Mannobiose phosphorylase Glycoside hydrolase family 130 Oligosaccharide synthesis Substrate specificity Structure-function relationship Enzyme-substrate interaction a b s t r a c t Glycoside hydrolase family 130 consists of phosphorylases and hydrolases for b-mannosides. Here, we characterized b-1,2-mannobiose phosphorylase from Listeria innocua (Lin0857) and determined its crystal structures complexed with b-1,2-linked mannooligosaccharides. b-1,2-Mannotriose was bound in a U-shape, interacting with a phosphate analog at both ends. Lin0857 has a unique dimer structure connected by a loop, and a significant open-close loop displacement was observed for substrate entry. A long loop, which is exclusively present in Lin0857, covers the active site to limit the pocket size. A structural basis for substrate recognition and phosphorolysis was provided.

Structural insights into the difference in substrate recognition of two mannoside phosphorylases from two GH130 subfamilies

“…According to the sequence‐based classification of glycoside hydrolases and related enzymes , both MGP and MOP were classified as members of the glycoside hydrolase family 130 (GH130), along with 1,4‐β‐mannosyl‐ N ‐acetylglucosamine phosphorylase (http://www.chem.qmul.ac.uk/iubmb/enzyme/EC2/4/1/320.html) , β‐1,4‐mannosyl‐[ N ‐glycan] phosphorylase , 1,2‐β‐oligomannan phosphorylase , β‐1,2‐mannobiose phosphorylase and β‐1,2‐mannosidase . GH130 was divided into three subfamilies (GH130_1, GH130_2 and GH130_NC) based on a phylogenetic analysis .…”

“…GH130_1 consists of MGPs, and GH130_2 consists of MOP, 1,4‐β‐mannosyl‐ N ‐acetylglucosamine phosphorylase and β‐1,4‐mannosyl‐[ N ‐glycan] phosphorylase. The phosphorylases and hydrolases those target β‐1,2 mannosidic linkages fall into GH130_NC . The three‐dimensional structures of several GH130 proteins are available, including Lin0857 (GH130_NC, phosphorolyzing β‐1,2‐mannosidic linkages) , BT3780 (GH130_NC, hydrolyzing β‐1,2‐mannosidic linkages) and some other uncharacterized proteins from GH130_2 and GH130_NC.…”

“…The phosphorylases and hydrolases those target β‐1,2 mannosidic linkages fall into GH130_NC . The three‐dimensional structures of several GH130 proteins are available, including Lin0857 (GH130_NC, phosphorolyzing β‐1,2‐mannosidic linkages) , BT3780 (GH130_NC, hydrolyzing β‐1,2‐mannosidic linkages) and some other uncharacterized proteins from GH130_2 and GH130_NC. Among these proteins, B. fragilis MGP ( Bf MGP) and β‐1,4‐mannosyl‐[ N ‐glycan] phosphorylase from an uncultivated Bacteroides bacteria (Uhgb_MP) were characterized, which belong to GH130_1 and GH130_2, respectively .…”

Mannoside recognition and degradation by bacteria

“…Finally, two GH130s from Dyadobacter fermentans DSM 18053 and Listeria innocua clip 11242 have been reported recently to exhibit β-1,2 mannosidase and β-1,2 mannoside phosphorylase (Tsuda et al, 2015) activities, respectively, but no biological function has yet been associated with these proteins.…”

Analysis of the diversity of the glycoside hydrolase family 130 in mammal gut microbiomes reveals a novel mannoside-phosphorylase function

“…The newly created GT108 family only contains six characterized members, all acting on mannogen, a linear β-1,2polymannoside. In contrast, the GH130 family is much more polyspecific, with 18 characterized members, of which 3 are β-1,2mannosidases [12,13] and 14 are glycoside phosphorylases acting on a large variety of substrates, such asβ-1,4-mannooligosaccharides [14][15][16][17][18], β-1,4mannosyl-N-acetyl-glucosamine [19],β-1,4mannosyl-N,N'-diacetylchitobiose [20],β-1,2-mannobiose [21,22],β-1,2-oligomannans [21] andβ-1,3-mannobiose [23]. Their large diversity of specificities, the tolerance of some enzymes towards acceptors, and their ability to generate αMan1P through phosphorolysis of cheap substrates for the synthesis of various heteromannosides, makes GH130 glycoside phosphorylases attractive enzymes for the in vitro synthesis of β-mannosidic linkages, which are considered to be some of the most challenging glycosidic linkages in synthetic carbohydrate chemistry [24].…”