A serine protease inhibitor from Musca domestica larva exhibits inhibitory activity against elastase and chymotrypsin

• Published in: Biotechnology letters Vol. 38; no. 7; pp. 1147 - 1153
• Main Authors: Tang, Yan, Wang, Ying, Pei, Zhihuan, Li, Wenting, Zhang, Dandan, Liu, Lei, Kong, Lingcong, Liu, Shuming, Jiang, Xiuyun, Ma, Hongxia
• Format: Journal Article
• Language: English
• Published: Dordrecht Springer Netherlands 01.07.2016; Springer Nature B.V
• Subjects: affinity chromatography; Animals; Applied Microbiology; Biochemistry; Biomedical and Life Sciences; Biotechnology; cDNA libraries; Chymotrypsin; Chymotrypsin - metabolism; complementary DNA; E coli; Elastase; Enzyme Activation - drug effects; Enzymes; Escherichia coli; Gene expression; gene expression regulation; Gene Library; Genes; Houseflies - metabolism; Insects; Larva - metabolism; Life Sciences; Microbiology; molecular cloning; Musca domestica; Nickel; Original Research Paper; Pancreatic Elastase - metabolism; papain; peptidase K; Protease inhibitors; Proteases; Proteinase inhibitors; Proteins; Recombinant; Serine Proteinase Inhibitors; trypsin; Elastase inhibitor; Gene cloning; Protein expression; Chymotrypsin inhibitor; Serine protease inhibitor; Serpins; Musca domestica
• ISSN: 0141-5492; 1573-6776
• DOI: 10.1007/s10529-016-2089-0

Objective Insect-derived serine protease inhibitors (serpins) exhibit multiple inhibitory activities, but so far, no functional roles for serpins of Musca domestica have been identified. Here, the functional features of M. domestica serine protease inhibitor (MDSPI16) were characterized. Results Hundred forty seven differentially expressed genes including the MDSPI16 gene were screened by constructing the subtractive cDNA library. The 1154-bp full-length MDSPI16 gene was cloned, and the recombinant MDSPI16 serpin protein was expressed as a 42.6 kDa protein in an Escherichia coli expression system. The recombinant MDSPI16 protein was purified using Ni–NTA affinity chromatography, and the inhibitory activity of MDSPI16 was assessed. MDSPI16 did not inhibit trypsin, papain, or proteinase K but strongly inhibited elastase (K i  = 2.8 nM) and chymotrypsin (K i  = 28 nM). The inhibitory activity of MDSPI16 remained stable over from 37 to 100 °C and from pH 2 to 12. Conclusions The MDSPI16 exhibited inhibitory activity against elastase and chymotrypsin and the inhibitory activity remained stable.