ABSTRACT
Features of gene expression of the secreted Bacillus pumilus metalloendopeptidase belonging to the adamalysin/reprolysin family were investigated. In the regulatory region of the gene, we identified hypothetical binding sites for transcription factors CcpA and TnrA. We found that the expression of the metalloendopeptidase gene is controlled by mechanisms of carbon and nitrogen catabolite repression. In experiments involving nitrogen metabolism regulatory protein mutant strains, we found that the control of the metalloendopeptidase gene expression involves proteins of ammonium transport GlnK and AmtB interacting with the TnrA-regulator.
Subject(s)
Bacillus pumilus/enzymology , Bacterial Proteins/biosynthesis , Gene Expression Regulation, Bacterial/physiology , Gene Expression Regulation, Enzymologic/physiology , Metalloendopeptidases/biosynthesis , Bacillus pumilus/genetics , Bacterial Proteins/genetics , Metalloendopeptidases/genetics , Nitrogen/metabolism , Transcription Factors/genetics , Transcription Factors/metabolismABSTRACT
Here wediscuss known properties of metzincin metalloproteinases, their structure, physiological roles in the cell and potential medical uses. We also present results describing a novel extracellular metzincin metalloproteinase from Bacillus pumilus with a unique combination of properties typical for both astacins and adamalysins.
Subject(s)
Bacillus/enzymology , Metalloproteases/chemistry , Protein Structure, Tertiary , Zinc/metabolism , Amino Acid Sequence , Humans , Metalloendopeptidases/chemistry , Metalloendopeptidases/metabolism , Metalloproteases/metabolism , Metalloproteases/therapeutic use , Protein ConformationABSTRACT
Heterologous gene expression of extracellular minor metalloendopeptidase of Bacillus pumilus 3-19 in protease-deficient B. subtilis strain has been studied. The fraction of enzyme in total pool of B. pumilus 3-19 secreted proteases composes less than 8%. The enzyme was isolated from culture liquid of recombinant strain, its primary structure was determined, physicochemical properties were investigated. It was concluded that secreted metallo endopeptidase of B. pumilus 3-19 represents the first prokaryotic homolog of eukaryotic adamalysin/reprolysin protein family.
Subject(s)
Bacillus/enzymology , Metalloendopeptidases/chemistry , Metalloendopeptidases/isolation & purification , Amino Acid Sequence , Enzyme Stability , Kinetics , Molecular Sequence Data , Prokaryotic Cells , Protein Structure, TertiaryABSTRACT
In this review the main families of endopeptidases belonging to the clan of metzincins of zinc-dependent metalloproteinases in organisms of wide evolutional range from bacteria to mammals are considered. The data on classification, physicochemical properties, substrate specificity, and structural features of this group of enzymes are given. The activation mechanisms of metzincins, the role of these proteins in organisms, and their participation in various physiological processes are discussed.
Subject(s)
Metalloendopeptidases/chemistry , Amino Acid Motifs , Binding Sites , Catalytic Domain , Metalloendopeptidases/classification , Metalloendopeptidases/metabolism , Zinc/chemistry , Zinc/metabolismABSTRACT
A novel zinc-dependent metalloendopeptidase of Bacillus intermedius (MprBi) was purified from the culture medium of a recombinant strain of Bacillus subtilis. The amino acid sequence of the homogeneous protein was determined using MALDI-TOF mass spectrometry. The sequence of the first ten residues from the N-terminus of the mature protein is ASTGSQKVTV. Physicochemical properties of the enzyme and its substrate specificity have been studied. The molecular weight of the metalloproteinase constitutes 19 kDa, the K(m) and k(cat) values are 0.06 mM and 1210 sec⻹, respectively, and the pI value is 5.4. The effect of different inhibitors and metal ions on the enzyme activity has been studied. Based on the analysis of the amino acid sequence of the active site motif and the Met-turn together with the enzyme characteristics, the novel bacterial metalloproteinase MprBi is identified as a metzincin clan adamalysin/reprolysin-like metalloprotease.
