Structural And Catalytic Insights Into A Novel Glycosyltransferase GlyE From Streptococcus Pneumoniae

The serine-rich repeat glycoproteins(SRRPs)that exclusively exist in Gram-positive bacteria are a growing family of surface adhesions SRRPs are involved in biofilm formation,bacterial adhesion and virulence.Streptococcus pneumoniae SRRP,termed PsrP forms the hair-like structure on the surface of cell wall.PsrP is highly glycosylated by a series of glycosyltransferases.Beyond the well-studied first two steps of the glycosylation of PsrP,the subsequent glycosylation steps remains unknown.Study of the structure and function of these glycosyltransferases will help us understand the glycosylation mechanism and biogenesis of PsrP,which might guide the reasonal design of novel drugs of antibiotics.We determined the crystal structures of apo-and UDP-bound forms of the glycosyltransferase GlyE at 2.4 A and 1.9 A respectively.Structural annlysis enabled us to identify that GlyE is a galactosyltransferase of GT8 family.Its N-terminal core domain adopts a typical metal-dependent GT-A fold.The C-terminal domain represents a single Rossmann-like fold(termed "add-on" domain)with unknown function.Further structural comparison of GlyE with its homologs suggests that this "add-on" domain might be involved in the recognition and catalysis of its acceptor PsrP.Taken together,the GlyE structure revealed a novel structure of GT-A glycosyltransferase with a fusion of the Rossmann-like fold "add-on" domain.Moreover,we tested the enzymatic activity of other glycosyltransferases involed in PsrP glycosylation,and identified their donor substrate.For example,GlyA.GlyB and GlyF can simultaneously hydrolyzed UDP-Gal and UDP-Glc,indicating that they may be branching enzymes.GlyG which contains a single GT2 domain could only hydrolyze UDP-Glc.These results will provide the basis for the further study of PsrP biogenesis.