Part:BBa_K5246019

HB HfsG Glycosyltransferase

Usage and Biology

The HfsG gene encodes a 331 amino acid protein, homologous to family 2 glycosyltransferases, in Hirschia baltica. This protein transfers sugar units from uridine diphosphate N-acetylglucosamine (UDP-GlcNAc) to the growing chain in synthesizing the holdfast polysaccharide repeat units in the cytoplasm. Studies have shown that HfsG mutants were completely devoid of any holdfast material.

This part also has a his-tagged variant BBa_K5246030.

Sequence and Features

Assembly Compatibility:

• 10
  COMPATIBLE WITH RFC[10]
• 12
  INCOMPATIBLE WITH RFC[12]

  Illegal NheI site found at 852
  
• 21
  INCOMPATIBLE WITH RFC[21]

  Illegal BglII site found at 747
  
• 23
  COMPATIBLE WITH RFC[23]
• 25
  COMPATIBLE WITH RFC[25]
• 1000
  COMPATIBLE WITH RFC[1000]

Experimental characterization

Bioinformatic analysis

CDD and protein BLAST analysis suggest that HfsG is a glucosyltransferase family 2 protein. Proteins of this family are involved in cell wall biosynthesis. HfsG is similar to the WecA protein in E.Coli that catalyzes the transfer of the GlcNAc-1-phosphate moiety from UDP-GlcNAc onto the carrier lipid undecaprenyl phosphate. In the case of HfsG, it catalyzes the transfer of UDP-GlcNAc to the sugar acceptor made earlier in the holdfast synthesis pathway.

Protein topology analysis using DeepTMHMM suggests that HfsG is a globular protein located in the cytoplasm. AlphaFold 3 structures further confirm it. A pTM score above 0.5 suggests that the predicted overall structure may closely resemble the true protein fold, while ipTM indicates the accuracy of the subunit positioning within the complex. Values higher than 0.8 represent confident, high-quality predictions.

HfsG is family 2 glycosyltransferase similar to WecA of E.Coli. This globular protein transfers UDP-GlcNAc to the acceptor molecule, our conclusions are in agreement with existing research. [1][2][3]

References

1. Toh, E., Kurtz, Harry D. and Brun, Y.V. (2008) ‘Characterization of the Caulobacter crescentus holdfast polysaccharide biosynthesis pathway reveals significant redundancy in the initiating glycosyltransferase and polymerase steps’, Journal of Bacteriology, 190(21), pp. 7219–7231. doi:10.1128/jb.01003-08.
2. Hardy, G.G. et al. (2018) ‘Mutations in sugar-nucleotide synthesis genes restore holdfast polysaccharide anchoring to Caulobacter crescentus holdfast anchor mutants’, Journal of Bacteriology, 200(3). doi:10.1128/jb.00597-17.
3. Sulkowski, N.I. et al. (2019) ‘A multiprotein complex anchors adhesive holdfast at the outer membrane of Caulobacter crescentus’, Journal of Bacteriology, 201(18). doi:10.1128/jb.00112-19.