Peptidase inhibitors from the salivary glands of the cockroach Nauphoeta cinerea

• Published in: Insect biochemistry and molecular biology 2009-12, Vol.39 (12), p.920-930
• Main Authors: Taranushenko, Yuliya, Vinokurov, Konstantin S., Kludkiewicz, Barbara, Kodrík, Dalibor, Sehnal, František
• Format: Article
• Language: English
• Subjects: Affinity chromatography; Amino Acid Sequence; amino acid sequences; Animals; Base Sequence; Blaberidae; Chymotrypsin; Cockroach digestion; Cockroaches - metabolism; complementary DNA; digestive physiology; enzyme activity; enzyme inactivation; enzyme inhibition; enzyme inhibitors; Gene Expression Regulation; insect genetics; insect pests; insect physiology; Insect saliva; Kazal inhibitors; molecular genetics; Molecular Sequence Data; Nauphoeta cinerea; nucleotide sequences; peptidases; Peptide Hydrolases - metabolism; Protease Inhibitors - metabolism; Proteinase K; saliva; salivary glands; sequence analysis; Subtilisin; trypsin
• ISSN: 0965-1748; 1879-0240
• DOI: 10.1016/j.ibmb.2009.11.002

Inhibitory activity against subtilisin, proteinase K, chymotrypsin and trypsin was detected in the salivary glands and saliva of the cockroach Nauphoeta cinerea (Blattoptera: Blaberidae). Fractionation of the salivary glands extract by affinity chromatography followed by reverse-phase HPLC yielded five subtilisin-inhibiting peptides with molecular masses ranging from 5 to 14 kDa. N-terminal sequences and subsequently full-length cDNAs of inhibitors designated NcPIa and NcPIb were obtained. The NcPIa cDNA contains 216 nucleotides and encodes a pre-peptide of 72 amino-acid residues of which 19 make up the signal peptide. The cDNA of NcPIb consists of 240 nucleotides and yields a putative secretory peptide of 80 amino-acid residues. Mature NcPIa (5906.6 Da, 53 residues) and NcPIb (6713.3 Da, 60 residues) are structurally similar (65.4% amino acid overlap) single-domain Kazal-type peptidase inhibitors. NcPIa with Arg in P1 position and typical Kazal motif VCGSD interacted stoichiometrically (1:1) with subtilisin and was slightly less active against proteinase K. NcPIb with Leu in P1 and modified Kazal motif ICGSD had similar activity on subtilisin and no on proteinase K but was active on chymotrypsin.