Further Characterization of Earthworm Serine Proteases: Cleavage Specificity Against Peptide Substrates and on Autolysis

1999 ◽  
Vol 63 (11) ◽  
pp. 2031-2033 ◽  
Author(s):  
Nobuyoshi NAKAJIMA ◽  
Manabu SUGIMOTO ◽  
Kohji ISHIHARA ◽  
Kaoru NAKAMURA ◽  
Hiroki HAMADA
Keyword(s):  
Serine Proteases ◽  
Peptide Substrates ◽  
Cleavage Specificity

Kinetic Characterization of Anticarsia gemmatalis Digestive Serine- Proteases and the Inhibitory Effect of Synthetic Peptides

2018 ◽  
Vol 24 (11) ◽  
Author(s):  
Adriana M. Patarroyo-Vargas ◽  
Yaremis B. Merino-Cabrera ◽  
Jose C. Zanuncio ◽  
Francelina Rocha ◽  
Wellington G. Campos ◽  
...  
Keyword(s):  
Synthetic Peptides ◽  
Serine Proteases ◽  
Inhibitory Effect ◽  
Anticarsia Gemmatalis ◽  
Kinetic Characterization

Cleavage Specificity of Human Rhinovirus-2 2A Protease for Peptide Substrates

1997 ◽  
Vol 235 (3) ◽  
pp. 562-566 ◽  
Author(s):  
Q.May Wang ◽  
Wolfgang Sommergruber ◽  
Robert B. Johnson
Keyword(s):  
Human Rhinovirus ◽  
Peptide Substrates ◽  
Cleavage Specificity ◽  
2A Protease

scholarly journals Recombinant Deg/HtrA proteases from Synechocystis sp. PCC 6803 differ in substrate specificity, biochemical characteristics and mechanism

2011 ◽  
Vol 435 (3) ◽  
pp. 733-742 ◽  
Author(s):  
Pitter F. Huesgen ◽  
Helder Miranda ◽  
XuanTam Lam ◽  
Manuela Perthold ◽  
Holger Schuhmann ◽  
...  
Keyword(s):  
Serine Proteases ◽  
Protein Quality ◽  
Biochemical Characterization ◽  
Pdz Domain ◽  
Control Mechanisms ◽  
Ph Optimum ◽  
Periplasmic Proteins ◽  
Synechocystis Sp ◽  
Pcc 6803

Cyanobacteria require efficient protein-quality-control mechanisms to survive under dynamic, often stressful, environmental conditions. It was reported that three serine proteases, HtrA (high temperature requirement A), HhoA (HtrA homologue A) and HhoB (HtrA homologue B), are important for survival of Synechocystis sp. PCC 6803 under high light and temperature stresses and might have redundant physiological functions. In the present paper, we show that all three proteases can degrade unfolded model substrates, but differ with respect to cleavage sites, temperature and pH optima. For recombinant HhoA, and to a lesser extent for HtrA, we observed an interesting shift in the pH optimum from slightly acidic to alkaline in the presence of Mg2+ and Ca2+ ions. All three proteases formed different homo-oligomeric complexes with and without substrate, implying mechanistic differences in comparison with each other and with the well-studied Escherichia coli orthologues DegP (degradation of periplasmic proteins P) and DegS. Deletion of the PDZ domain decreased, but did not abolish, the proteolytic activity of all three proteases, and prevented substrate-induced formation of complexes higher than trimers by HtrA and HhoA. In summary, biochemical characterization of HtrA, HhoA and HhoB lays the foundation for a better understanding of their overlapping, but not completely redundant, stress-resistance functions in Synechocystis sp. PCC 6803.

scholarly journals Arginine Inhibits Serpins

2007 ◽  
Vol 13 (2) ◽  
pp. 213-218
Author(s):  
Thomas W. Stief
Keyword(s):  
Esterase Activity ◽  
Serine Proteases ◽  
Biological Fluid ◽  
Microtiter Plate ◽  
Human Albumin ◽  
Amidolytic Activity ◽  
Peptide Substrates ◽  
Plate Reader ◽  
High Concentrations ◽  
Very High

Serine protease inactivators (serpins) are important regulators in biochemistry. Often it is necessary to block the serpin action, that is, to stabilize the sample. The guanidine group of arginine is the ligand for the active center pocket of many serine proteases. Arginine or guanidine inhibits serine proteases, and arginine belongs to the reactive P1-P1' center of many serpins. The plasmatic antithrombin, antiplasmin, or anti-C1-esterase activity was determined: A total of 20 µL of pooled normal plasma or 7% human albumin was added to 100 µL of 0—2.67 M arginine, pH 8.6, 10 µL of 26 mIU/mL thrombin in 7% human albumin, and 30 µL of 1.7 mM CHG-Ala-Arg-pNA (37°C). ΔA at 405 nm was determined, by using a microtiter plate reader. Thrombin was substituted by plasmin or C1-esterase, and the chromogenic peptide substrates <Glu-Phe-Lys-pNA or MeOC-Lys(eCBO)-Gly-Arg-pNA, respectively, were used. The IC50 of arginine against plasmatic antithrombin activity is 580 mM; the IC 25 is 440 mM. The IC25 of arginine against plasmatic α 2-antiplasmin or C1-inactivator is 1650 mM. The amidolytic activity of thrombin, plasmin, and C1-esterase is inhibited similarly by arginine: the IC50 for arginine against the amidolytic activity of these proteases is about 400 mM. Arginine at very high concentrations inhibits serpins. This is important, if stabilization of a biological fluid is a prerequisite for valid activities of serine proteases. In addition, these high concentrations of arginine might be a new gentle principle to inhibit pathogens that need serpins for their pathophysiology.

Characterization of three alternative splice variants associated with the Arabidopsis rhomboid protein gene At1g74130

Botany ◽  
2013 ◽  
Vol 91 (12) ◽  
pp. 840-849 ◽  
Author(s):  
Joshua Powles ◽  
Katharine Sedivy-Haley ◽  
Eric Chapman ◽  
Kenton Ko
Keyword(s):  
Alternative Splicing ◽  
Alternative Splice ◽  
Splice Variant ◽  
Serine Proteases ◽  
Transmembrane Domain ◽  
Splice Variants ◽  
Genes Encoding ◽  
Different Characteristics ◽  
Alternative Splice Variants

Rhomboid serine proteases are grouped into three main types — secretases, presenilin-like associated rhomboid-like (PARL) proteases, and “inactive” rhomboid proteins. Although the three rhomboid groups are distinct, the different types are likely to operate within the same cell or compartment, such as observed in the plastids of Arabidopsis. There are four distinct plastid rhomboid genes at play in Arabidopsis plastids, two for active types (At1g25290 and At5g25752) and two for inactive forms (At1g74130 and At1g74140). The number of working plastid rhomboids is further increased by alternative splicing, as reported for At1g25290. To understand how the plastid rhomboid system works, it is necessary to identify all rhomboid forms in play. To this end, this study was designed to examine the alternative splicing activities of At1g74130, one of the two genes encoding proteolytically “inactive” plastid rhomboids. The exon mapping and DNA sequencing results obtained here indicate the presence of three prominent alternative splice variants in the At1g74130 transcript population. The dominant splice variant, L, encodes the full-length protein. The other two splice variants, M and S, produce proteins lacking sections from the carboxyl transmembrane domain region. The splice variants M and S appear to be at levels with functional potential and appear to adjust relative to each other during development and in response to changes in the level of Tic40, a component of the plastid translocon. The splice variant proteins themselves exhibit different characteristics with respect to rhomboid protein–substrate interactions. These differences were observed in bacterial co-expression pull-down assays and in yeast mitochondrial studies. When considered together, the data suggest that the alternative splicing of At1g74130 bears functional significance in Arabidopsis and is likely to be part of a mechanism for diversifying plastid rhomboid function.

Molecular characterization of a partial sequence encoding a novel Schistosoma mansoni serine protease

Parasitology ◽  
1997 ◽  
Vol 115 (4) ◽  
pp. 395-402 ◽  
Author(s):  
C. COCUDE ◽  
C. PIERROT ◽  
C. CETRE ◽  
C. GODIN ◽  
A. CAPRON ◽  
...  
Keyword(s):  
Schistosoma Mansoni ◽  
Serine Protease ◽  
Serine Proteases ◽  
Mrna Level ◽  
Cross Reactivity ◽  
Reading Frame ◽  
Consensus Sequences ◽  
Degenerate Oligonucleotide ◽  
Sequence Encoding

A PCR strategy using degenerate oligonucleotide primers based upon consensus sequences of the active site of serine proteases yielded a 467 bp fragment from genomic DNA from Schistosoma mansoni cercariae. The sequence presented a continuous open reading frame and the deduced amino acid sequence (156 aa) presented homologies with various serine proteases, in particular the highest percentage identity was observed with a mammalian plasma kallikrein. The expression of this serine protease was studied first at the mRNA level and it was only detected by RT-PCR in cercariae and in adult worms. At the protein level we were able to detect it by Western blotting and by using antigen extracts from metabolically radio-isotope labelled worms. The absence of any positive signal in Northern blot and the detection of the protein suggest that the mRNA has a very short half-life, however the protein may be accumulated in the parasite. The significance of identity with mammalian kallikrein was confirmed by cross-immunoreactivity with a native porcine pancreatic kallikrein. However, no cross-reactivity was observed with S. mansoni elastase, another serine protease. Thus, we suggest that the serine protease described in this paper is a kallikrein-like protease.

Study of the Proteolytic Activity of the Tropical Legume Crotalaria spectabilis

2012 ◽  
Vol 67 (9-10) ◽  
pp. 495-509 ◽  
Author(s):  
Juliana da Silva Pacheco ◽  
Raquel Elisa da Silva-Lopez
Keyword(s):  
Proteolytic Activity ◽  
Serine Proteases ◽  
Ph Value ◽  
Cysteine Proteases ◽  
Leaf Extracts ◽  
Serine Protease Activity ◽  
Proteolytic Activities ◽  
Low Levels ◽  
Crotalaria Spectabilis

The characterization of legume proteases contributes to the understanding of the physiology of plants and their interaction with the environment. Thirteen extracts from various parts of Crotalaria spectabilis were made using different extraction systems. The highest protein content was found in seeds, and the most pronounced proteolytic activity was observed in leaf extracts, with an optimal pH value in the alkaline range. Proteases in extracts from roots, stems, and fl owers were active in various pH ranges. Proteases in all extracts were maximally active between 30 °C and 60 °C and were thermostable (24 h, 60 °C). Hemoglobin, bovine serum albumin, casein, and gelatin were hydrolyzed by C. spectabilis extracts in different ways. The highest serine protease activity was found in leaves. Seeds contained high levels of serine proteases and low levels of cysteine proteases. Flowers, roots, and stems contained different levels of serine, aspartic, and metalloproteases, respectively. The proteolytic activities in extracts were modulated by cations and oxidants to various degrees. C. spectabilis proteases are differentially expressed in distinctive organs, and their stability against heat and oxidants makes this plant an important source of stable proteases

scholarly journals Characterization of two family AA9 LPMOs from Aspergillus tamarii with distinct activities on xyloglucan reveals structural differences linked to cleavage specificity

PLoS ONE ◽  
2020 ◽  
Vol 15 (7) ◽  
pp. e0235642 ◽  
Author(s):  
Antonielle V. Monclaro ◽  
Dejan M. Petrović ◽  
Gabriel S. C. Alves ◽  
Marcos M. C. Costa ◽  
Glaucia E. O. Midorikawa ◽  
...  
Keyword(s):  
Aspergillus Tamarii ◽  
Cleavage Specificity ◽  
Structural Differences

scholarly journals Cleavage Specificity ofMycobacterium tuberculosisClpP1P2 Protease and Identification of Novel Peptide Substrates and Boronate Inhibitors with Anti-bacterial Activity

2015 ◽  
Vol 290 (17) ◽  
pp. 11008-11020 ◽  
Author(s):  
Tatos Akopian ◽  
Olga Kandror ◽  
Christopher Tsu ◽  
Jack H. Lai ◽  
Wengen Wu ◽  
...  
Keyword(s):  
Bacterial Activity ◽  
Peptide Substrates ◽  
Cleavage Specificity
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