Mutant endoglycoceramidases with enhanced synthetic activity

A method of producing monosialoganglioside 1 (GM1), the method comprising:

• contacting a fluorinated monosialoganglioside 1 (GM1) sugar donor substrate and an acceptor substrate with a variant of a wild-type endoglycoceramidase in a reaction mixture, wherein the variant catalyzes the transfer of the saccharide moiety from the fluorinated GM1 donor substrate to the acceptor substrate at a rate that exceeds hydrolysis of the GM1, wherein said wild-type endoglycoceramidase has the amino acid sequence of SEQ. ID. NO.:
• 2 and said variant has a serine substitution of the Glu residue at position 351 of SEQ. ID. NO.:
• 2, and wherein the acceptor substrate is sphingosine or a sphingosine analog.

A method of producing a glycolipid, the method comprising:

• contacting a donor substrate having an activated saccharide moiety and an acceptor substrate with a variant of a wild-type endoglycoceramidase in a reaction mixture, wherein the variant catalyzes the transfer of the saccharide moiety from the donor substrate to the acceptor substrate at a rate that exceeds hydrolysis of the glycolipid, wherein said variant comprises an amino acid substitution of the Glu residue 351 within SEQ ID NO:2 selected from E351S, E351G, and E351A and wherein the acceptor substrate is sphingosine or a sphingosine analog.