Characterisation of endoglucanases EGB and EGC from Fibrobacter succinogenes - Archive ouverte HAL Access content directly
Journal Articles Biochimica et Biophysica Acta (BBA) - Protein Structure and Molecular Enzymology Year : 2000

Characterisation of endoglucanases EGB and EGC from Fibrobacter succinogenes

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Véronique Broussolle
  • Function : Author
Peter Pristas
  • Function : Author
Jean-Pierre Girardeau
  • Function : Author
Geneviève Gaudet
  • Function : Author
Evelyne Forano
  • Function : Author

Abstract

The enzymatic properties of two endoglucanases from Fibrobacter succinogenes, EGB and EGC, were analysed. EGB and EGC were purified from recombinant Escherichia coli cultures expressing their gene. The failure of purification of EGB by classical techniques led us to produce antipeptide antibodies that allowed immunopurification of the protein from E. coli as well as its detection in F. succinogenes cultures. Synthetic peptides were selected from the predicted primary structure of EGB, linked to bovine serum albumin and used as immunogens to obtain specific antibodies. One of the polyclonal antipeptide antisera was used to purify EGB. EGC was purified by affinity chromatography with Ni-NTA resin. The endo mode of action of the two enzymes on carboxymethyl-cellulose was different. The values of K(m) and V(max) were respectively 13.6 mg/ml and 46 micromol/min mg protein for EGB, and 7 mg/ml and 110 micromol/min mg protein for EGC. The reactivity of the antipeptide and the anti-EGC sera with F. succinogenes proteins of molecular mass different from that of EGB and EGC produced in E. coli suggested post-translational modification of the two enzymes in F. succinogenes cultures. Expression of endB and endC genes in F. succinogenes was confirmed by RT-PCR.

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hal-03802697 , version 1 (06-10-2022)

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Christel Béra-Maillet, Véronique Broussolle, Peter Pristas, Jean-Pierre Girardeau, Geneviève Gaudet, et al.. Characterisation of endoglucanases EGB and EGC from Fibrobacter succinogenes. Biochimica et Biophysica Acta (BBA) - Protein Structure and Molecular Enzymology, 2000, 1476 (2), pp. 191-202. ⟨10.1016/s0167-4838(99)00255-1⟩. ⟨hal-03802697⟩
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