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Cloning of Feather Degrading Minor Extracellular Protease from Bacillus cereus DCUW: Dissection of the Structural Domains

¹ University of Calcutta, MPI, Marburg;
² University of Calcutta

ABSTRACT

Bacterial extracellular proteases play an important role in cell survival and cell-cell communication. A high molecular weight minor extracellular protease (Vpr) from a feather degrading bacterium, Bacillus cereus DCUW, has been reported by our laboratory. In the present study, we have cloned and expressed Vpr in Escherichia coli. Complete nucleotide sequencing of this gene predicted that the protease is a member of serine protease family and SMART domain analysis revealed that the protease consists of an N-terminal signal sequence for secretion; a subtilisin_N sequence that signatures for N-terminal processing; a catalytic S_8 peptidase domain; and finally a long C-terminal protease associated (PA) region containing nine intrinsically disordered sub-domains. Four truncated constructs of the minor extracellular protease (Vpr) are cloned and expressed in E.coli. We found that the catalytic domain (amino acids residues,172-583) is sufficient to show protease activity. Maturation of the Vpr protease needed both N-terminal and C-terminal processing. We have demonstrated that the oligomerization property is associated with the C-terminal protease associated domain and also shown that the substrate binding specificity to raw feather resides in this domain. This is the first report on structure-function studies of minor extracellular protease (Vpr) from B.cereus DCUW.

³ E-mail: djcbcg{at}caluniv.ac.in