Cloning and characterisation of the UDP-glucose 4'-epimerase of Trypanosoma cruzi.
Trypanosoma cruzi incorporates galactose into many of its cell-surface glycoconjugates but it is unable to transport this sugar through its hexose transporter. Epimerisation of UDP-glucose to UDP-galactose by UDP-glucose 4'-epimerase may be the only way that the parasites can obtain galactose. Here, we describe cloning the T. cruzi UDP-Glc 4'-epimerase (TcGALE) gene and show that it is functional by complementing an Escherichia coli epimerase-deficient strain. The T. cruzi GALE gene encodes a 42.4 kDa protein and the recombinant protein expressed in E. coli is a homodimer in solution with a specific activity of 3.8 U mg(-1) and K(m) for UDP-Gal of 114 microM. Unlike the human epimerase, T. cruzi UDP-Glc 4'-epimerase is unable to inter-convert UDP-N-acetylglucosamine and UDP-N-acetylgalactosamine. This may explain why T. cruzi initiates O-glycosylation of its abundant GPI-anchored surface mucins via GlcNAcalpha1-O-Thr/Ser rather than the GalNAcalpha1-O-Thr/Ser linkage that is common for mucins from many other eukaryotes.
Roper
Janine R
JR
Division of Biological Chemistry and Molecular Microbiology, The School of Life Sciences, University of Dundee, Dundee DD1 5EH, Scotland, UK.
Ferguson
Michael A J
MA
D000595:Amino Acid Sequence; D000818:Animals; D003001:Cloning, Molecular; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D005690:Galactose / Q000378:metabolism; D005816:Genetic Complementation Test; D006031:Glycosylation; D006801:Humans; D008969:Molecular Sequence Data; D051381:Rats; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D014349:Trypanosoma cruzi / Q000201:enzymology* / Q000235:genetics; D014534:UDPglucose 4-Epimerase / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*
D011994:Recombinant Proteins; D014534:UDPglucose 4-Epimerase; D005690:Galactose