%0 Journal Article
%T Human gut microbes express functionally distinct endoglycosidases to metabolize the same N-glycan substrate.
%A Sastre DE
%A Sultana N
%A V A S Navarro M
%A Huliciak M
%A Du J
%A Cifuente JO
%A Flowers M
%A Liu X
%A Lollar P
%A Trastoy B
%A Guerin ME
%A Sundberg EJ
%J Nat Commun
%V 15
%N 1
%D 2024 Jun 15
%M 38879612
%F 17.694
%R 10.1038/s41467-024-48802-3
%X Bacteroidales (syn. Bacteroidetes) are prominent members of the human gastrointestinal ecosystem mainly due to their efficient glycan-degrading machinery, organized into gene clusters known as polysaccharide utilization loci (PULs). A single PUL was reported for catabolism of high-mannose (HM) N-glycan glyco-polypeptides in the gut symbiont Bacteroides thetaiotaomicron, encoding a surface endo-β-N-acetylglucosaminidase (ENGase), BT3987. Here, we discover an ENGase from the GH18 family in B. thetaiotaomicron, BT1285, encoded in a distinct PUL with its own repertoire of proteins for catabolism of the same HM N-glycan substrate as that of BT3987. We employ X-ray crystallography, electron microscopy, mass spectrometry-based activity measurements, alanine scanning mutagenesis and a broad range of biophysical methods to comprehensively define the molecular mechanism by which BT1285 recognizes and hydrolyzes HM N-glycans, revealing that the stabilities and activities of BT1285 and BT3987 were optimal in markedly different conditions. BT1285 exhibits significantly higher affinity and faster hydrolysis of poorly accessible HM N-glycans than does BT3987. We also find that two HM-processing endoglycosidases from the human gut-resident Alistipes finegoldii display condition-specific functional properties. Altogether, our data suggest that human gut microbes employ evolutionary strategies to express distinct ENGases in order to optimally metabolize the same N-glycan substrate in the gastroinstestinal tract.