Structural basis for catalysis and inhibition of N-glycan processing class I alpha 1,2-mannosidases.
Endoplasmic reticulum (ER) class I alpha1,2-mannosidase (also known as ER alpha-mannosidase I) is a critical enzyme in the maturation of N-linked oligosaccharides and ER-associated degradation. Trimming of a single mannose residue acts as a signal to target misfolded glycoproteins for degradation by the proteasome. Crystal structures of the catalytic domain of human ER class I alpha1,2-mannosidase have been determined both in the presence and absence of the potent inhibitors kifunensine and 1-deoxymannojirimycin. Both inhibitors bind to the protein at the bottom of the active-site cavity, with the essential calcium ion coordinating the O-2' and O-3' hydroxyls and stabilizing the six-membered rings of both inhibitors in a (1)C(4) conformation. This is the first direct evidence of the role of the calcium ion. The lack of major conformational changes upon inhibitor binding and structural comparisons with the yeast alpha1, 2-mannosidase enzyme-product complex suggest that this class of inverting enzymes has a novel catalytic mechanism. The structures also provide insight into the specificity of this class of enzymes and provide a blueprint for the future design of novel inhibitors that prevent degradation of misfolded proteins in genetic diseases.
Vallee
F
F
Program in Structural Biology and Biochemistry, Research Institute, Hospital for Sick Children, Toronto, Ontario M5G 1X8, Canada.
Karaveg
K
K
Herscovics
A
A
Moremen
K W
KW
Howell
P L
PL
D017485:1-Deoxynojirimycin / Q000737:chemistry / Q000378:metabolism; D000470:Alkaloids / Q000737:chemistry / Q000378:metabolism; D000595:Amino Acid Sequence; D000818:Animals; D002118:Calcium / Q000378:metabolism; D002384:Catalysis; D004721:Endoplasmic Reticulum / Q000201:enzymology*; D006801:Humans; D008361:Mannosidases / Q000037:antagonists & inhibitors*; D008969:Molecular Sequence Data; D011134:Polysaccharides / Q000378:metabolism; D011487:Protein Conformation; D011817:Rabbits; D011994:Recombinant Proteins / Q000302:isolation & purification; D013329:Structure-Activity Relationship
D000470:Alkaloids; D011134:Polysaccharides; D011994:Recombinant Proteins; C065629:kifunensine; D017485:1-Deoxynojirimycin; D008361:Mannosidases; C046090:mannosyl-oligosaccharide 1,2-alpha-mannosidase; D002118:Calcium