scholarly journals Amino acid sequence around the thiol and reactive acyl groups of human complement component C4

1981 ◽  
Vol 199 (2) ◽  
pp. 359-370 ◽  
Author(s):  
R D Campbell ◽  
J Gagnon ◽  
R R Porter
Keyword(s):  
Thiol Group ◽  
Ester Bond ◽  
Human Complement ◽  
Active Molecule ◽  
Free Thiol ◽  
Continuous Sequence ◽  
N Terminus ◽  
Free Thiol Group ◽  
Fourth Component ◽  
Human Complement Component

Activation of the fourth component of complement (C4) by C1s results in the generation of a reactive acyl group, able to react with putrescine, and in the release of a free thiol group that cannot be detected in the native haemolytically active molecule. Both the reactive acyl group and the free thiol group have been shown to reside in C4d, a fragment of the alpha′-chain of C4b derived from digestion of the molecule with the control proteins C3b inactivator and C4-binding protein. Peptides derived from CNBr digestion of [1,4-14C]putrescine-labelled and iodo(2-14C]acetic acid-labelled C4d have been obtained and used to establish a continuous sequence of 88 residues from the N-terminus of the molecule. The thiol and reactive acyl groups are contained in an octapeptide that shows near identity with the equivalent sequences reported for alpha 2-macroglobulin and C3. Other adjacent short sections also show homology of sequence between the three proteins, and it is highly likely that they contribute to the overall structure that gives a unique reactivity to the thiol ester bond postulated to exist in the native forms of the three proteins.

scholarly journals The reaction of iodine and thiol-blocking reagents with human complement components C2 and factor B. Purification and N-terminal amino acid sequence of a peptide from C2a containing a free thiol group

1983 ◽  
Vol 213 (1) ◽  
pp. 201-209 ◽  
Author(s):  
C Parkes ◽  
J Gagnon ◽  
M A Kerr
Keyword(s):  
Thiol Group ◽  
Haemolytic Activity ◽  
Human Complement ◽  
Ph Optimum ◽  
Complement Components ◽  
Factor B ◽  
Molecule Reaction ◽  
Free Thiol ◽  
Disulphide Bond Formation ◽  
Free Thiol Group

Human complement components C2 and Factor B each contain one free thiol group/molecule. Reaction with p-chloromercuribenzoate destroyed the haemolytic activity of C2 but had no effect on Factor B. Reaction of C2 with I2 gave a 16-fold enhancement of its haemolytic activity. The pH optimum for the reaction was 7.0. The I2 reacted at the thiol group in C2 with a stoicheiometry of 1 mol of I2/mol of C2. The product of the reaction was unaffected by millimolar concentrations of dithiothreitol; however, azide and cyanide were inhibitory. Reaction with azide did not result in re-expression of the thiol group. Mild oxidation of the thiol group with m-chloroperbenzoic acid did not enhance the haemolytic activity. The results suggest that reaction with I2 causes intramolecular covalent, but not disulphide, bond formation. I2 reacted with Factor B at the free thiol group without affecting the haemolytic activity. A CNBr-cleavage peptide from C2a (obtained by cleavage of C2 by subcomponent C1s) containing the free thiol group was isolated. Automated Edman degradation of the peptide showed that it was the N-terminal peptide of C2a. The free thiol group was identified at position 18.

scholarly journals 1250. Novel Boronic Acid Transition State Analogs (BATSI) with in vitro inhibitory activity against class A, B and C β-lactamases

2020 ◽  
Vol 7 (Supplement_1) ◽  
pp. S643-S643
Author(s):  
Maria F Mojica ◽  
Christopher Bethel ◽  
Emilia Caselli ◽  
Magdalena A Taracila ◽  
Fabio Prati ◽  
...  
Keyword(s):  
Active Site ◽  
Inhibitory Activity ◽  
Covalent Bond ◽  
Thiol Group ◽  
Viable Cell ◽  
Free Thiol ◽  
Transition State Analogs ◽  
Cell Counts ◽  
Free Thiol Group

Abstract Background Catalytic mechanisms of serine β-lactamases (SBL; classes A, C and D) and metallo-β-lactamases (MBLs) have directed divergent strategies towards inhibitor design. SBL inhibitors act as high affinity substrates that -as in BATSIs- form a reversible, dative covalent bond with the conserved active site Ser. MBL inhibitors bind the active-site Zn2+ ions and displace the nucleophilic OH-. Herein, we explore the efficacy of a series of BATSI compounds with a free-thiol group at inhibiting both SBL and MBL. Methods Exploratory compounds were synthesized using stereoselective homologation of (+) pinandiol boronates to introduce the amino group on the boron-bearing carbon atom, which was subsequently acylated with mercaptopropanoic acid. Representative SBL (KPC-2, ADC-7, PDC-3 and OXA-23) and MBL (IMP-1, NDM-1 and VIM-2) were purified and used for the kinetic characterization of the BATSIs. In vitro activity was evaluated by a modified time-kill curve assay, using SBL and MBL-producing strains. Results Kinetic assays revealed that IC50 values ranged from 1.3 µM to >100 µM for this series. The best compound, s08033, demonstrated inhibitory activity against KPC-2, VIM-2, ADC-7 and PDC-3, with IC50 in the low μM range. Reduction of at least 1.5 log10-fold of viable cell counts upon exposure to sub-lethal concentrations of antibiotics (AB) + s08033, compared to the cells exposed to AB alone, demonstrated the microbiological activity of this novel compound against SBL- and MBL-producing E. coli (Table 1). Table 1 Conclusion Addition of a free-thiol group to the BATSI scaffold increases the range of these compounds resulting in a broad-spectrum inhibitor toward clinically important carbapenemases and cephalosporinases. Disclosures Robert A. Bonomo, MD, Entasis, Merck, Venatorx (Research Grant or Support)

Fatty acids binding to human serum albumin: Changes of reactivity and glycation level of Cysteine-34 free thiol group with methylglyoxal

2014 ◽  
Vol 224 ◽  
pp. 42-50 ◽  
Author(s):  
Ivan D. Pavićević ◽  
Vesna B. Jovanović ◽  
Marija M. Takić ◽  
Ana Z. Penezić ◽  
Jelena M. Aćimović ◽  
...  
Keyword(s):  
Fatty Acids ◽  
Human Serum Albumin ◽  
Serum Albumin ◽  
Human Serum ◽  
Thiol Group ◽  
Free Thiol ◽  
Free Thiol Group

Post-proline endopeptidase. Interaction of the enzyme with substrates containing disulfide and thioether bond

1986 ◽  
Vol 51 (1) ◽  
pp. 234-240 ◽  
Author(s):  
Karel Hauzer ◽  
Tomislav Barth ◽  
Linda Hauzerová ◽  
Jana Barthová ◽  
Pavel Hrbas ◽  
...  
Keyword(s):  
Arginine Vasopressin ◽  
Thiol Group ◽  
Disulfide Bridge ◽  
Complex Interaction ◽  
Enzyme Inactivation ◽  
Molar Ratio ◽  
Disulfide Exchange ◽  
Neurohypophysial Hormones ◽  
Free Thiol ◽  
Free Thiol Group

The free thiol group of post-proline endopeptidase (EC 3.4.21.26) can interact with the disulfide bridge contained in some of the substrates of this enzyme (neurohypophysial hormones and some of their analogues). The influence of these interactions on the activity of this enzyme was studied using several substances modelling individual types of interactions: thiol-disulfide exchange, catalytic interaction and a complex interaction including the two preceding types. Deamino-1-carba-oxytocin is catalytically hydrolysed in the concentration range up to 10-3mol/l, oxytocin and arginine-vasopressin are catalytically hydrolysed in concentrations of 10-5 to 10-8 mol/l. A reaction leading to inactivation of the enzyme prevails at concentrations of 10-3 to 10-4 mol/l. When inactivated by lower concentrations of arginine-vasopressin (up to a molar ratio of 1 : 1), the enzyme can be reactivated by incubation with dithiothreitol, higher concentrations of arginine-vasopresson cause irreversible enzyme inactivation.

Photo-triggered generation of a free thiol group on DNA: application to DNA conjugation

2012 ◽  
Vol 53 (1) ◽  
pp. 78-81 ◽  
Author(s):  
Tadao Takada ◽  
Yuta Kawano ◽  
Mitsunobu Nakamura ◽  
Kazushige Yamana
Keyword(s):  
Thiol Group ◽  
Free Thiol ◽  
Free Thiol Group

scholarly journals Free Thiol Group of MD-2 as the Target for Inhibition of the Lipopolysaccharide-induced Cell Activation

2009 ◽  
Vol 284 (29) ◽  
pp. 19493-19500 ◽  
Author(s):  
Mateja Manček-Keber ◽  
Helena Gradišar ◽  
Melania Iñigo Pestaña ◽  
Guillermo Martinez de Tejada ◽  
Roman Jerala
Keyword(s):  
Cell Activation ◽  
Thiol Group ◽  
Free Thiol ◽  
Free Thiol Group

scholarly journals Mutagenic structure/function analysis of the cytoplasmic cysteines of the insulin receptor

1995 ◽  
Vol 306 (3) ◽  
pp. 811-820 ◽  
Author(s):  
S L Macaulay ◽  
M Polites ◽  
M J Frenkel ◽  
D R Hewish ◽  
C W Ward
Keyword(s):  
Cell Lines ◽  
Kinase Activity ◽  
Mutant Cell ◽  
Thiol Group ◽  
Insulin Binding ◽  
Beta Subunit ◽  
Wild Type ◽  
Free Thiol ◽  
Free Thiol Group ◽  
Mutant Receptors

Native human insulin receptor (hIR) has been reported to contain only one free thiol group proposed to lie near the ATP-binding. domain of its beta-subunit [Finn, Ridge and Hofmann (1990) Proc. Natl. Acad. Sci. U.S.A. 87, 419-423]. The present study investigated the role of the six cytoplasmic cysteines of the beta-subunit of the hIR using a mutagenic approach in which insulin receptors, mutated at each cytoplasmic cysteine (to alanine) in turn, were transfected into Chinese hamster ovary (CHO) cells. Cell lines expressing hIR mutation at high level were obtained which, by both flow-cytometric analysis towards an hIR-specific monoclonal antibody (83-7) and insulin-binding analysis, were similar to the well-characterized CHOT cell line which overexpresses native hIR. The ED50 and Kd values of the mutant receptors were the same as those of the wild-type hIR. Each of the mutant receptors signalled insulin action to stimulate receptor autophosphorylation and kinase activity as well as glucose utilization to levels appropriate for the receptor level expressed. In contrast, insulin-stimulated thymidine uptake and glucose-transport responses of two of the six mutant cell lines, those expressing Cys981Ala and Cys1245Ala, were impaired compared with that of the native hIR-expressing cell line, CHOT. The beta-subunits of each of the hIR cytoplasmic cysteine mutant cell lines could be alkylated specifically with N-[3H]ethylmaleimide. The kinase activity of each receptor was inhibited by N-ethylmaleimide and stimulated by iodoacetamide, indicating that none of the cytoplasmic cysteines alone contributes the single free thiol group to the hIR structure. We conclude that the cytoplasmic cysteines of the hIR have a predominantly passive role in hIR activity although Cys-981 and Cys-1245 do affect mitogenic and glucose-transport responses of the receptor. Our findings indicate that the stoicheiometry of a single free thiol group/mol of insulin-binding activity noted in previous studies is either spread fractionally over a number of the cytoplasmic cysteines or is one of the four cysteines in the ectodomain of the hIR beta-subunit. Alternatively, the mutagenesis performed in the present study may enable differential exposure of a second titratable cysteine in wild-type and mutant receptors.

scholarly journals Cysteine Analogs with a Free Thiol Group Promote Fertilization by Reducing Disulfide Bonds in the Zona Pellucida of Mice1

2015 ◽  
Vol 92 (4) ◽  
Author(s):  
Toru Takeo ◽  
Yuka Horikoshi ◽  
Satohiro Nakao ◽  
Kazuhito Sakoh ◽  
Yuta Ishizuka ◽  
...  
Keyword(s):  
Zona Pellucida ◽  
Disulfide Bonds ◽  
Thiol Group ◽  
Free Thiol ◽  
Free Thiol Group
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