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Partial characterization of trypsin-like protease and molecular cloning of a trypsin-like precursor cDNA in salivary glands of Lygus lineolaris.
Comp Biochem Physiol B Biochem Mol Biol. 2002 Mar; 131(3):453-63.CB

Abstract

Based on substrate specificity, an alkaline pH optimum, sensitivity to selected proteinase inhibitors, and molecular analysis, we provide evidence for the presence of a trypsin-like serine proteinase in the salivary gland complex (SGC) of the tarnished plant bug, Lygus lineolaris (Palisot de Beauvois) (Heteroptera: Miridae). The predominant activity in extracts of the SGC against N(2)-benzoyl-L-arginine-p-nitroanilide (L-BApNA) was at pH 10, but a minor peak of activity also occurred at pH 5. The major BApNAase activity focused at 10.4 during preparative isoelectric focusing and was eluted with an apparent molecular weight of 23,000 from a calibrated gel filtration column. The BApNAase fraction gave a single major band when analyzed on a casein zymogram. The activity was completely suppressed by the serine protease inhibitors, phenylmethylsulfonyl fluoride (PMSF) and lima bean trypsin inhibitor. A cDNA coding for a trypsin-like protein in the salivary glands of L. lineolaris was cloned and sequenced. The 971bp cDNA contained an 873-nucleotide open reading frame encoding a 291-amino acid trypsin precursor. The encoded protein included amino acid sequence motifs that are conserved with four homologous serine proteases from other insects. Typical features of the putative trypsin-like protein from L. lineolaris included the serine protease active site (His(89), Asp(139), Ser(229)), conserved cysteine residues for disulfide bridges, the residues (Asp(223), Gly(252), Gly(262)) that determine trypsin specificity, and both zymogen signal and activation peptides. Cloning and sequencing of a trypsin-like precursor cDNA provided additional direct evidence for trypsin like enzymes in the salivary glands of L. lineolaris.

Authors+Show Affiliations

Biological Control and Mass Rearing Research Unit, USDA/ARS, P.O. Box 5367, Mississippi State, MS 39762, USA. fzeng@bcmrru.msstate.eduNo affiliation info availableNo affiliation info available

Pub Type(s)

Journal Article

Language

eng

PubMed ID

11959027

Citation

Zeng, F, et al. "Partial Characterization of Trypsin-like Protease and Molecular Cloning of a Trypsin-like Precursor cDNA in Salivary Glands of Lygus Lineolaris." Comparative Biochemistry and Physiology. Part B, Biochemistry & Molecular Biology, vol. 131, no. 3, 2002, pp. 453-63.
Zeng F, Zhu Y, Cohen A. Partial characterization of trypsin-like protease and molecular cloning of a trypsin-like precursor cDNA in salivary glands of Lygus lineolaris. Comp Biochem Physiol B Biochem Mol Biol. 2002;131(3):453-63.
Zeng, F., Zhu, Y., & Cohen, A. (2002). Partial characterization of trypsin-like protease and molecular cloning of a trypsin-like precursor cDNA in salivary glands of Lygus lineolaris. Comparative Biochemistry and Physiology. Part B, Biochemistry & Molecular Biology, 131(3), 453-63.
Zeng F, Zhu Y, Cohen A. Partial Characterization of Trypsin-like Protease and Molecular Cloning of a Trypsin-like Precursor cDNA in Salivary Glands of Lygus Lineolaris. Comp Biochem Physiol B Biochem Mol Biol. 2002;131(3):453-63. PubMed PMID: 11959027.
* Article titles in AMA citation format should be in sentence-case
TY - JOUR T1 - Partial characterization of trypsin-like protease and molecular cloning of a trypsin-like precursor cDNA in salivary glands of Lygus lineolaris. AU - Zeng,F, AU - Zhu,Y, AU - Cohen,A, PY - 2002/4/18/pubmed PY - 2002/9/11/medline PY - 2002/4/18/entrez SP - 453 EP - 63 JF - Comparative biochemistry and physiology. Part B, Biochemistry & molecular biology JO - Comp Biochem Physiol B Biochem Mol Biol VL - 131 IS - 3 N2 - Based on substrate specificity, an alkaline pH optimum, sensitivity to selected proteinase inhibitors, and molecular analysis, we provide evidence for the presence of a trypsin-like serine proteinase in the salivary gland complex (SGC) of the tarnished plant bug, Lygus lineolaris (Palisot de Beauvois) (Heteroptera: Miridae). The predominant activity in extracts of the SGC against N(2)-benzoyl-L-arginine-p-nitroanilide (L-BApNA) was at pH 10, but a minor peak of activity also occurred at pH 5. The major BApNAase activity focused at 10.4 during preparative isoelectric focusing and was eluted with an apparent molecular weight of 23,000 from a calibrated gel filtration column. The BApNAase fraction gave a single major band when analyzed on a casein zymogram. The activity was completely suppressed by the serine protease inhibitors, phenylmethylsulfonyl fluoride (PMSF) and lima bean trypsin inhibitor. A cDNA coding for a trypsin-like protein in the salivary glands of L. lineolaris was cloned and sequenced. The 971bp cDNA contained an 873-nucleotide open reading frame encoding a 291-amino acid trypsin precursor. The encoded protein included amino acid sequence motifs that are conserved with four homologous serine proteases from other insects. Typical features of the putative trypsin-like protein from L. lineolaris included the serine protease active site (His(89), Asp(139), Ser(229)), conserved cysteine residues for disulfide bridges, the residues (Asp(223), Gly(252), Gly(262)) that determine trypsin specificity, and both zymogen signal and activation peptides. Cloning and sequencing of a trypsin-like precursor cDNA provided additional direct evidence for trypsin like enzymes in the salivary glands of L. lineolaris. SN - 1096-4959 UR - https://www.unboundmedicine.com/medline/citation/11959027/Partial_characterization_of_trypsin_like_protease_and_molecular_cloning_of_a_trypsin_like_precursor_cDNA_in_salivary_glands_of_Lygus_lineolaris_ L2 - https://linkinghub.elsevier.com/retrieve/pii/S1096495901005140 DB - PRIME DP - Unbound Medicine ER -