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Trypsin-like Proteinases (trypsin-like + proteinase)

Distribution by Scientific Domains
Life Sciences57%
Medical Sciences42%

Selected Abstracts

cDNA sequence, mRNA expression and genomic DNA of trypsinogen from the Indianmeal moth, Plodia interpunctella

INSECT MOLECULAR BIOLOGY, Issue 1 2000
Y. C. Zhu
Abstract Trypsin-like enzymes are major insect gut enzymes that digest dietary proteins and proteolytically activate insecticidal proteins produced by the bacterium Bacillus thuringiensis (Bt). Resistance to Bt in a strain of the Indianmeal moth, Plodia interpunctella, was linked to the absence of a major trypsin-like proteinase (Oppert et al., 1997). In this study, trypsin-like proteinases, cDNA sequences, mRNA expression levels and genomic DNAs from Bt-susceptible and -resistant strains of the Indianmeal moth were compared. Proteinase activity blots of gut extracts indicated that the susceptible strain had two major trypsin-like proteinases, whereas the resistant strain had only one. Several trypsinogen-like cDNA clones were isolated and sequenced from cDNA libraries of both strains using a probe deduced from a conserved sequence for a serine proteinase active site. cDNAs of 852 nucleotides from the susceptible strain and 848 nucleotides from the resistant strain contained an open reading frame of 783 nucleotides which encoded a 261-amino acid trypsinogen-like protein. There was a single silent nucleotide difference between the two cDNAs in the open reading frame and the predicted amino acid sequence from the cDNA clones was most similar to sequences of trypsin-like proteinases from the spruce budworm, Choristoneura fumiferana, and the tobacco hornworm, Manduca sexta. The encoded protein included amino acid sequence motifs of serine proteinase active sites, conserved cysteine residues, and both zymogen activation and signal peptides. Northern blotting analysis showed no major difference between the two strains in mRNA expression in fourth-instar larvae, indicating that transcription was similar in the strains. Southern blotting analysis revealed that the restriction sites for the trypsinogen genes from the susceptible and resistant strains were different. Based on an enzyme size comparison, the cDNA isolated in this study corresponded to the gene for the smaller of two trypsin-like proteinases, which is found in both the Bt-susceptible and -resistant strains of the Indianmeal moth. The sequences reported in this paper have been deposited in the GenBank database (accession numbers AF064525 for the RC688 strain and AF064526 for HD198). [source]

Gingival crevicular fluid laminin-5 ,2-chain levels in periodontal disease

JOURNAL OF CLINICAL PERIODONTOLOGY, Issue 7 2006
Gülnur Emingil
Abstract Aim: Our study aimed to examine the molecular forms and gingival crevicular fluid (GCF) levels of laminin-5 ,2-chain in patients with different periodontal disease, and compare the effects of P.gingivalis trypsin-like proteinase on intact laminin-5 ,2-chain species. Methods: Eighteen patients with generalized aggressive periodontitis (G-AgP), 29 patients with chronic periodontitis (CP), 20 with gingivitis and 20 periodontally healthy subjects were included. Probing depth, clinical attachment loss, presence of bleeding on probing and plaque were recorded. Molecular forms and GCF laminin-5 ,2-chain levels and the effects of P. gingivalis trypsin-like proteinase on intact laminin-5 ,2-chain were analysed by computer-quantitated Western immunoblotting. Results: Laminin-5 ,2-chain 40 and 70 kDa fragments could be detected in all groups, in varying levels. The CP group had elevated GCF laminin-5 ,2-chain fragment levels compared with the gingivitis and healthy groups (p<0.008). The G-AgP group had GCF laminin-5 ,2-chain fragment levels similar to the gingivitis and healthy groups (p>0.008). GCF laminin-5 ,2-chain fragments differed clearly from the multiple lower molecular size fragments of P.gingivalis trypsin-laminin-5 ,2-chain proteinases. Conclusion: Increased GCF laminin-5 ,2-chain fragments in periodontitis sites with deep periodontal pocket suggest that these cleaved 40 and 70 kDa fragments could reflect the extent of the inflammatory reaction in CP. [source]

cDNA sequence, mRNA expression and genomic DNA of trypsinogen from the Indianmeal moth, Plodia interpunctella

INSECT MOLECULAR BIOLOGY, Issue 1 2000
Y. C. Zhu
Abstract Trypsin-like enzymes are major insect gut enzymes that digest dietary proteins and proteolytically activate insecticidal proteins produced by the bacterium Bacillus thuringiensis (Bt). Resistance to Bt in a strain of the Indianmeal moth, Plodia interpunctella, was linked to the absence of a major trypsin-like proteinase (Oppert et al., 1997). In this study, trypsin-like proteinases, cDNA sequences, mRNA expression levels and genomic DNAs from Bt-susceptible and -resistant strains of the Indianmeal moth were compared. Proteinase activity blots of gut extracts indicated that the susceptible strain had two major trypsin-like proteinases, whereas the resistant strain had only one. Several trypsinogen-like cDNA clones were isolated and sequenced from cDNA libraries of both strains using a probe deduced from a conserved sequence for a serine proteinase active site. cDNAs of 852 nucleotides from the susceptible strain and 848 nucleotides from the resistant strain contained an open reading frame of 783 nucleotides which encoded a 261-amino acid trypsinogen-like protein. There was a single silent nucleotide difference between the two cDNAs in the open reading frame and the predicted amino acid sequence from the cDNA clones was most similar to sequences of trypsin-like proteinases from the spruce budworm, Choristoneura fumiferana, and the tobacco hornworm, Manduca sexta. The encoded protein included amino acid sequence motifs of serine proteinase active sites, conserved cysteine residues, and both zymogen activation and signal peptides. Northern blotting analysis showed no major difference between the two strains in mRNA expression in fourth-instar larvae, indicating that transcription was similar in the strains. Southern blotting analysis revealed that the restriction sites for the trypsinogen genes from the susceptible and resistant strains were different. Based on an enzyme size comparison, the cDNA isolated in this study corresponded to the gene for the smaller of two trypsin-like proteinases, which is found in both the Bt-susceptible and -resistant strains of the Indianmeal moth. The sequences reported in this paper have been deposited in the GenBank database (accession numbers AF064525 for the RC688 strain and AF064526 for HD198). [source]

Momordica charantia trypsin inhibitor II inhibits growth and development of Helicoverpa armigera

INSECT SCIENCE, Issue 5 2009
Manasi Alok Telang
Abstract, Bitter gourd (Momordica charantia L.) seeds contain several squash-type serine proteinase inhibitors (PIs), which inhibit the digestive proteinases of the polyphagous insect pest Helicoverpa armigera. In the present work isolation of a DNA sequence encoding the mature peptide of a trypsin inhibitor McTI-II, its cloning and expression as a recombinant protein using Pichia pastoris have been reported. Recombinant McTI-II inhibited bovine trypsin at 1: 1 molar ratio, as expected, but did not inhibit chymotrypsin or elastase. McTI-II also strongly inhibited trypsin-like proteinases (81% inhibition) as well as the total proteolytic activity of digestive proteinases (70% inhibition) from the midgut of H. armigera larvae. The insect larvae fed with McTI-II-incorporated artificial diet suffered over 70% reduction in the average larval weight after 12 days of feeding. Moreover, ingestion of McTI-II resulted in 23% mortality in the larval population. The strong antimetabolic activity of McTI-II toward H. armigera indicates its probable use in developing insect tolerance in susceptible plants. [source]

Selective inhibition of Porphyromonas gingivalis growth by a factor Xa inhibitor, DX-9065a

JOURNAL OF PERIODONTAL RESEARCH, Issue 3 2006
Kenji Matsushita
Background:,Porphyromonas gingivalis is a causative bacterium of adult periodontitis. However, there is no drug specific for P. gingivalis and for its virulence factor. Objectives:, The objective of this study was to examine the effects of a new selective inhibitor of activated factor X, DX-9065a, on growth of Porphyromonas gingivalis and other periodontopathic bacteria. Methods:, We incubated P. gingivalis and other periodontopathic bacteria in the presence or absence of DX-9065a and examined the effect of DX-9065a on bacterial growth and trypsin-like activity in its cultures. We also examined the effects of DX9065a on amidolytic activity of purified trypsin-like proteinases (gingipains RgpA and RgpB), from P. gingivalis and on trypsin-like activity in gingival crevicular fluids from patients with adult periodontitis. Results:, DX-9065a selectively inhibited the growth of P. gingivalis and Prevotella intermedia, and its effect on P. gingivalis was bactericidal. Trypsin-like proteinase activity was detected in P. gingivalis, and the activity was strongly inhibited by DX-9065a. DX-9065a even inhibited amidolytic activity of RgpA and RgpB from P. gingivalis. Furthermore, trypsin-like proteinase activity in gingival crevicular fluids was strongly inhibited by DX-9065a. Conclusions:, DX-9065a inhibits P. gingivalis growth in part through to its ability to inhibit the trypsin-like proteinase activity in P. gingivalis and may be useful for a new drug for treatment of adult periodontitis. [source]