Detail of GM0009


Summary

The genetically modified enzybiotic, named CHAPSH3b , constructed by Domains Assembly strategy, improve the lytic activity.
With  CHAPSH3b
  Sequence Length:  259 AA.
  Mass:  29220.7 Da.
  Isoelectric Point:  10.21
  Function:  increased CHAP activity against both, S. aureus Sa9 and S. aureus MRSA N315 strains, for 64-fold

Construction

GM0009, constructed by Domains Assembly strategy.
    No schema construted on GM0009
Details:
GM0009[1-156]: derive from protein B7T0E8 sequence 475-634

B7T0E8, 634 AA., the Gp58 from Staphylococcus phage phiSauS-IPLA88

Source: Staphylococcus phage phiSauS-IPLA88

Domains and repeats

15-149:  IPR007921, the CHAP domain
483-629:  IPR013338, the Lysozyme domain, subfamily 2

GO term prediction

Biological Process:  0009253, the peptidoglycan catabolic process
Biological Process:  0044036, the cell wall macromolecule metabolic process
Molecular Function:  0004040, the amidase activity?
Molecular Function:  0016787, the hydrolase activity?


Linking to UniprotKB
Linging to InterPro
GM0009[156-261]: derive from protein P10547 sequence 411-485

P10547, 493 AA., the Lysostaphin from Staphylococcus simulans

Source: Staphylococcus simulans

Domains and repeats

239-382:  IPR011055, the Duplicated hybrid motif?
411-481:  IPR003646, the SH3-like domain, bacterial-type
278-367:  IPR003646, the Peptidase M23?

GO term prediction

    No GO info found on GM0009

Linking to UniprotKB
Linging to InterPro

Annotation

1-156
156-261

15-149
: EAD, CHAP domain, derived from phage vB_SauS-phiIPLA88 (phiIPLA88)PG hydrolase(HydH5);
156-261
: CBD, SH3b domain, derived from lysostaphin;
Domain and repeats:
156-257
: IPR003646,SH3-like domain, bacterial-type;
9-151
: IPR013338,Lysozyme domain, subfamily 2;
GO term prediction:
0016787, the hydrolase activity
MGLPNPKDRKPTASEVVEWALYMAKNRRVIDVDRSYGGQCWDVPNYILERYWGFRTWGNANAMAQKSNYRGRDFKIYRNTASFTPKPGDWAVWANRNPGHVAIVVGPADKNAFVSVDQNWYTANWSGSPPYKIKHTYHDGPGGVTHFVRPPYHPDK-GGTVTPTPNTGWKTNKYGTLYKSESASFTPNTDIITRTTGPFRSMPQSGVLKAGQTIHYDEVMKQDGHVWVGYTGNSGQRIYLPVRTWNKSTNTLGVLWGTIK
  BlastP to GMEnzy

Production

  1.  Expressed by pET24a in E. coli BL21(DE3),  Purified by NiNTA nickel column chromatography (Qiagen, Valencia, CA).

Activity

  1.  lytic activity test on Staphylococcus aureus (live S. aureus Sa9 cells ) by the turbidity-reduction assays
  showed specific activity of 0.109 ∆OD600nm min-1µM-1

Reference

  1. Lorena Rodriguez-Rubio, Beatriz Martinez, Ana Rodriguez1, David M. Donovan, and Pilar Garc¨ªa. (2012) Enhanced staphylolytic activity of the Staphylococcus aureus bacteriophage vB_SauS-phiIPLA88 HydH5 virion associated peptidoglycan hydrolase: fusions, deletions and synergy with LysH5. Appl. Environ. Microbiol.. 4431-2:32-41. [doi:10.1128/AEM.07621-11] [PMID:22267667] [FULL TEXT]
  2. Rodriguez-Rubio, L.Martinez, B.Donovan, D. M.Garcia, P.Rodriguez, A.. (2013) Potential of the virion-associated peptidoglycan hydrolase HydH5 and its derivative fusion proteins in milk biopreservation. PLoS One. 01:e54828. [doi:10.1371/journal.pone.0054828] [PMID:23359813] [FULL TEXT]

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