Part:BBa_K5246001
CB2/CB2A HfsA Part of export protein complex
Introduction
Usage and Biology
Gene HfsA from Caulobacter crescentus encodes a protein of 502 amino acids from complex that in combination with HfsB is responsible for controlled polymerisation of holdfast pollysaccharide. HfsA is a polysaccharide copolymerase Wzz
Sequence and Features
- 10COMPATIBLE WITH RFC[10]
- 12INCOMPATIBLE WITH RFC[12]Illegal NotI site found at 388
- 21COMPATIBLE WITH RFC[21]
- 23COMPATIBLE WITH RFC[23]
- 25INCOMPATIBLE WITH RFC[25]Illegal NgoMIV site found at 64
Illegal NgoMIV site found at 256
Illegal NgoMIV site found at 265
Illegal NgoMIV site found at 837
Illegal NgoMIV site found at 1000 - 1000COMPATIBLE WITH RFC[1000]
Experimental characterization
Bioinformatic analysis
Conservative Domain Database analysis revealed that the HfsA protein contains domains characteristic of the GumC superfamily, which typically function in exopolysaccharide export within the cell wall or membrane. Additionally, it shows significant similarity to the PEP-CTERM superfamily, proteins that are generally involved in determining polysaccharide chain length.
NCBI protein BLAST analysis revealed significant similarities between HfsA and a capsular polysaccharide biosynthesis protein from the ABC transporter family of Caldimonas thermodepolymerans. Using the DeepTMHMM tool to analyze its transmembrane structure, it was predicted that HfsA spans the membrane twice, embedding itself firmly in the cell envelope.
With AlphaFold3, we assessed different configurations of HfsA subunits, focusing on ipTM and pTM scores. 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 We hypothesise that HfsA transmembrane protein consists of 8 subunits with confidence scores of ipTM = 0.72 and pTM = 0.73 (Fig.1).
Based on this data, we hypothesize that HfsA is a transmembrane protein responsible for exporting polysaccharides from the cell. Similar findings were proposed by earlier research. [1][2][3][4]
, 
Fig. 1. Alphafold3 structure showing assembled hfsA protein of 8 subunits
[[Image:Figure 1: Fig. 1. Alphafold3 structure showing assembled hfsA protein of 8 subunits]
References
1. Kurtz, H.D. and Smith, J. (1994) ‘The Caulobacter crescentushold fast: Identification of holdfast attachment complex genes’, FEMS Microbiology Letters, 116(2), pp. 175–182. doi:10.1111/j.1574-6968.1994.tb06697.x.
2. Javens, J. et al. (2013) ‘Bypassing the need for subcellular localization of a polysaccharide export‐anchor complex by overexpressing its protein subunits’, Molecular Microbiology, 89(2), pp. 350–371. doi:10.1111/mmi.12281.
3. Smith, C.S. et al. (2003) ‘Identification of genes required for synthesis of the adhesive holdfast in caulobacter crescentus’, Journal of Bacteriology, 185(4), pp. 1432–1442. doi:10.1128/jb.185.4.1432-1442.2003.
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