Biochemical and molecular characterization of a novel metalloprotease from Pseudomonas fluorescens strain TBS09.

A novel extracellular protease called MPDZ was purified and characterized from Pseudomonas fluorescens strain TBS09. The enzymatic properties of MPDZ were investigated using biochemical and biophysical methods. Matrix assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF/MS) analysis revealed that it was a monomer with a molecular mass of 50013.17Da. The NH-terminal 27 amino acid sequence of MPDZ showed high homology with those of Pseudomonas-proteases of the serralysin family. MPDZ showed optimal activity at pH 7 and 60°C. It was totally inhibited by EGTA, EDTA, and 1,10-phenanthroline, suggesting its belonging to the metalloprotease family. Because of the interesting properties, the mpDZ gene encoding MPDZ was cloned, sequenced, and expressed in E. coli. The deduced amino acid sequence showed a strong homology with other Pseudomonas-metalloproteases. The highest sequence identity value (97%) was obtained with AprX from P. fluorescens strain CY091, with only 12 different amino acid residues. The physico-chemical properties of the extracellular purified recombinant enzyme (rMPDZ) were similar to those of MPDZ. Overall, MPDZ is bestowed with a number of promising biochemical properties that might give new opportunities for its biocatalytic applications. These data constitute an essential first step towards an understanding of the properties of MPDZ enzyme.