Trypanosoma cruzi trans-sialidase operates through a covalent sialyl-enzyme intermediate: tyrosine is the catalytic nucleophile.
Modified sialic acid substrates have been used to label Trypanosoma cruzi trans-sialidase, demonstrating that the enzyme catalyses the transfer of sialic acid through a covalent glycosyl-enzyme intermediate, a mechanism common to most retaining glycosidases. Peptic digestion of labeled protein, followed by LC-MS/MS analysis of the digest, identified Tyr342 as the catalytic nucleophile. This is the first such example of a retaining glycosidase utilizing an aryl glycoside intermediate. It is suggested that this alternative choice of nucleophile is a consequence of the chemical nature of sialic acid. A Tyr/Glu couple is invoked to relay charge from a remote glutamic acid, thereby avoiding electrostatic repulsion with the sialic acid carboxylate group.
Watts
Andrew G
AG
Department of Chemistry, University of British Columbia, 2036 Main Mall, Vancouver, B.C., Canada V6T 1Z1.
Damager
Iben
I
Amaya
Maria L
ML
Buschiazzo
Alejandro
A
Alzari
Pedro
P
Frasch
Alberto C
AC
Withers
Stephen G
SG
D000818:Animals; D002384:Catalysis; D004789:Enzyme Activation; D006023:Glycoproteins / Q000737:chemistry / Q000378:metabolism*; D007700:Kinetics; D009439:Neuraminidase / Q000737:chemistry / Q000378:metabolism*; D021241:Spectrometry, Mass, Electrospray Ionization; D014349:Trypanosoma cruzi / Q000201:enzymology*; D014443:Tyrosine / Q000737:chemistry / Q000378:metabolism*
D006023:Glycoproteins; D014443:Tyrosine; C069715:trans-sialidase; D009439:Neuraminidase