Purification and Characterization of the Lon-Like Recombinant Protease of Lactobacillus Plantarum IIA-1A5 Olfa Mega a*), Cece Sumantri b), Irma Isnafia Arief b), Cahyo Budiman b)
a) Faculty of Animal Science, University of Jambi
* olfa_mega[at]unja.ac.id
2) Department of Animal Production and Technology, Faculty of Animal Science, IPB University
Abstract
Lon protease is an enzyme responsible for degrading proteins. Lon-like proteases have the same effec as Lon. The gene encoding the Lon-like IIA-A5 was obtained from the genome of the bacterium, Lactobacillus plantarum IIA-1A5. The Lon-like IIA1A5 gene was cloned into the pET-28a (+) vector and expressed in Escherichia coli BL21(DE3) competent cells. The L. plantarum IIA-1A5 is a native Indonesian strain and has proteolytic activity in meat, processed meat products and casein. The study aims to purify and verify the proteolytic activity of Lon-like proteases produced by L plantarum IIA-1A5 in degrading animal product proteins. We used HisTrap SP FF 5 mL column with different imidazole concentrations for purification, namely 10- 50- 100- 150- 200- 250- 300- 350 and 500 mM. Then, the purified Lon-like IIA-1A5 was tested for optimal pH and temperature, and its ability to the hydrolyzed animal product proteins. The results showed that the specific activity, purity level, and yield of the purified Lon-like IIA-1A5 were 828.96 AU/mg, 2.2 times, and 224% respectively. The molecular weight of this protein was confirmed by SDS-PAGE of 43.76 kDa. The proteolytic activity of the enzyme is optimal at pH 9 and a temperature of 60 oC. Lon-like IIA-1A5 can generally degrade protein animal products. However, the enzyme can hydrolyze gelatin, myofibrillar protein, collagen, and sarcoplasmic protein better than casein and skim milk.
