Open AccessExpression induction of Serratia plymuthica UBCF_13 metalloprotease gene with various types of metal ions
Author(s) -
R. Zulfitri,
Siti Aisyah,
Maythesya Oktavioni,
R Fatiah,
J Jamsari
Publication year - 2021
Publication title -
iop conference series. earth and environmental science
Language(s) - English
Resource type - Journals
eISSN - 1755-1307
pISSN - 1755-1315
DOI - 10.1088/1755-1315/741/1/012039
Subject(s) - metalloproteinase , chemistry , proteases , metal ions in aqueous solution , biochemistry , gene expression , escherichia coli , plasmid , enzyme , microbiology and biotechnology , gene , biology , metal , organic chemistry
Bacteria as biocontrol agents can produce hydrolytic enzymes, one of them is a protease which plays an important role to degrade cell membranes or plasmalemma in fungi composed of protein. Metalloproteases are proteases that contain one or two metal ions on their active sites. The role of metal ions in metalloproteases is to activate water molecules, which act as nucleophiles in the catalysis process. The metalloprotease function of Serratia plymuthica UBCF_13 is unknown, therefore this study aimed to determine the effective metal ion to induce the expression of the UBCF_13 metalloprotease gene. The metalloprotease gene from Serratia plymuthica UBCF_13 was ligated into a pGEM T-easy plasmid. The recombinant plasmid was transformed into Escherichia coli BL21 by the heat shock method. The metalloprotease gene expression was induced by IPTG and a combination of several metal ions namely Zn 2+ , Mn 2+ , Fe 2+ , and Ca 2+ . The highest gene expression was characterized by the thickest protein band based on the result of SDS-PAGE visualization with a molecular weight of 27 kDa after being induced by Fe 2+ metal ions. Proteolytic activity of metalloprotease UBCF_13 was tested using solid LB media containing 2% skim milk. The longest clear zone diameter was obtained up to 0,83 cm after being induced by IPTG combined with Ca 2+ metal ion. These results can be used as a reference for the expression of the UBCF_13 metalloprotease gene. However, further enzyme purification is to be able the enzyme as an antifungal compound.