6027-13-0Relevant articles and documents
Functional consequences of homocysteinylation of the elastic fiber proteins fibrillin-1 and tropoelastin
Hubmacher, Dirk,Cirulis, Judith T.,Miao, Ming,Keeley, Fred W.,Reinhardt, Dieter P.
, p. 1188 - 1198 (2010)
Homocystinuria caused by cystathionine-β-synthase deficiency represents a severe form of homocysteinemias, which generally result in various degrees of elevated plasma homocysteine levels. Marfan syndrome is caused by mutations in fibrillin-1, which is one of the major constituents of connective tissue microfibrils. Despite the fundamentally different origins, both diseases share common clinical symptoms in the connective tissue such as long bone overgrowth, scoliosis, and ectopia lentis, whereas they differ in others. Fibrillin-1 contains ～13% cysteine residues and can be modified by homocysteine. We report here that homocysteinylation affects functional properties of fibrillin-1 and tropoelastin. We used recombinant fragments spanning the entire fibrillin-1 molecule to demonstrate that homocysteinylation, but not cysteinylation leads to abnormal self-interaction, which was attributed to a reduced amount of multimerization of the fibrillin-1 C terminus. The deposition of the fibrillin-1 network by human dermal fibroblasts was greatly reduced by homocysteine, but not by cysteine. Furthermore, homocysteinylation, but not cysteinylation of elastin-like polypeptides resulted in modified coacervation properties. In summary, the results provide new insights into pathogenetic mechanisms potentially involved in cystathionine-β-synthase-deficient homocystinuria.
Exploration of the six tryptophan residues of Escherichia coli cystathionine β-lyase as probes of enzyme conformational change
Jaworski, Allison F.,Aitken, Susan M.
, p. 138 - 144 (2013)
Cystathionine β-lyase (CBL) catalyzes the hydrolysis of l-cystathionine (l-Cth), producing l-homocysteine (l-Hcys), pyruvate and ammonia, in the second step of the transsulfuration pathway of bacteria and plants. A series of 17 site-directed variants of Escherichia coli CBL (eCBL) was constructed to probe the contributions of the six tryptophan residues (W131, W188, W230, W276, W300 and W340) to the fluorescence spectrum of eCBL and to assess their mutability and utility as conformational probes. The effects of these Trp → Phe substitutions on kcat and Km l-Cth are less than 2-fold, with the exception of the 8-fold increase in Kml-Cth observed for eCBL-W340F. The midpoint of thermal denaturation, as monitored by circular dichroism spectroscopy, is reduced 4.7 C by the W188F substitution while the targeted replacement of the other five tryptophans alter Tm by less than 1.7 C. The fluorescence spectrum of eCBL is dominated by W230 and the contribution of W340, situated in the active site, is minor. The observed 5-fold increase in the 336 nm fluorescence emission of W188 between 0 and 2 M urea, suggests a conformational change at the domain interface. Residues W188 and W340, conserved in proteobacterial CBL enzymes, are situated at the core of the domain interface that forms the active-site cleft. The results of this study suggest that W188 is a useful probe of subtle conformational changes at the domain interface and active site.
Identification of O-acetylhomoserine sulfhydrylase, a putative enzyme responsible for methionine biosynthesis in Clostridioides difficile: Gene cloning and biochemical characterizations
Kulikova, Vitalia V.,Revtovich, Svetlana V.,Bazhulina, Natalia P.,Anufrieva, Natalya V.,Kotlov, Mikhail I.,Koval, Vasiliy S.,Morozova, Elena A.,Hayashi, Hideyuki,Belyi, Yury F.,Demidkina, Tatyana V.
, p. 1815 - 1823 (2019)
O-acetylhomoserine sulfhydrylase (OAHS) is a pyridoxal 5′-phosphate-dependent enzyme involved in microbial methionine biosynthesis. In this study, we report gene cloning, protein purification, and some biochemical characteristics of OAHS from Clostridioides difficile. The enzyme is a tetramer with molecular weight of 185 kDa. It possesses a high activity in the reaction of L-homocysteine synthesis, comparable to reported activities of OAHSes from other sources. OAHS activity is inhibited by metabolic end product L-methionine. L-Propargylglycine was found to be a suicide inhibitor of the enzyme. Substrate analogue Nγ-acetyl-L-2,4-diaminobutyric acid is a competitive inhibitor of OAHS with Ki = 0.04 mM. Analysis of C. difficile genome allows to suggest that the bacterium uses the way of direct sulfhydrylation for the synthesis of L-methionine. The data obtained may provide the basis for further study of the role of OAHS in the pathogenic bacterium and the development of potential inhibitors.
Hyperhomocysteinemia leads to exacerbation of ischemic brain damage: Role of GluN2A NMDA receptors
Jindal, Ankur,Rajagopal, Sathyanarayanan,Winter, Lucas,Paul, Surojit,Poddar, Ranjana,Brigman, Jonathan,Allan, Andrea M.,Miller, Joshua W.,Jacobsen, Donald W.
, p. 287 - 302 (2019)
Hyperhomocysteinemia has been implicated in several neurodegenerative disorders including ischemic stroke. However, the pathological consequences of ischemic insult in individuals predisposed to hyperhomocysteinemia and the associated etiology are unknown. In this study, we evaluated the outcome of transient ischemic stroke in a rodent model of hyperhomocysteinemia, developed by subcutaneous implantation of osmotic pumps containing L-homocysteine into male Wistar rats. Our findings show a 42.3% mortality rate in hyperhomocysteinemic rats as compared to 7.7% in control rats. Magnetic resonance imaging of the brain in the surviving rats shows that mild hyperhomocysteinemia leads to exacerbation of ischemic injury within 24 h, which remains elevated over time. Behavioral studies further demonstrate significant deficit in sensorimotor functions in hyperhomocysteinemic rats compared to control rats. Using pharmacological inhibitors targeting the NMDAR subtypes, the study further demonstrates that inhibition of GluN2A-containing NMDARs significantly reduces ischemic brain damage in hyperhomocysteinemic rats but not in control rats, indicating that hyperhomocysteinemia-mediated exacerbation of ischemic brain injury involves GluN2A-NMDAR signaling. Complementary studies in GluN2A-knockout mice show that in the absence of GluN2A-NMDARs, hyperhomocysteinemia-associated exacerbation of ischemic brain injury is blocked, confirming that GluN2A-NMDAR activation is a critical determinant of the severity of ischemic damage under hyperhomocysteinemic conditions. Furthermore, at the molecular level we observe GluN2A-NMDAR dependent sustained increase in ERK MAPK phosphorylation under hyperhomocysteinemic condition that has been shown to be involved in homocysteine-induced neurotoxicity. Taken together, the findings show that hyperhomocysteinemia triggers a unique signaling pathway that in conjunction with ischemia-induced pathways enhance the pathology of stroke under hyperhomocysteinemic conditions.
COMPOSITONS AND METHODS FOR TREATING BRAIN INJURY
-
Page/Page column 42, (2020/02/06)
A method for treating a hyperhomocysteinemic subject having cerebral ischemic stroke generally includes administering to the hyperhomocysteinemic subject, following cerebral stroke, a composition that includes an inhibitor or an antagonist of a GluN2A-containing N-methyl-D-aspartate receptor (NMDAR) in an amount effective to ameliorate at least one symptom or clinical sign of cerebral stroke.
Preparation method of N-fluorenylmethoxycarbonyl-S-(4-methoxytribenzyl)-L-homocysteine
-
Paragraph 0010; 0021; 0025; 0029; 0033, (2019/10/04)
The invention relates to a preparation method of N-fluorenylmethoxycarbonyl-S-(4-methoxytribenzyl)-L-homocysteine, and solves the technical problem that there is no effective synthesis method in the prior art. The preparation method comprises following steps: (1) dissolving L-methionine into a solvent, and under the action of a strong reducing agent, removing one methyl group to prepare a crude product of L-homocysteine; (2) dissolving the crude product of L-homocysteine and 4-methoxytriphenyl chloromethane into a solvent, slowly adding a strong acid, TLC monitoring the reactions for 12 to 24 hours, carrying out suction filtration, adjusting the pH to 7 by an alkaline solution, carrying out suction filtration, and washing the obtained solid by a solvent to obtain S-(4-methoxytribenzyl)-L-homocysteine; and (3) dissolving S-(4-methoxytribenzyl)-L-homocysteine into a solvent, adding a fluorenylmethoxycarbonyl protective group onto S-(4-methoxytribenzyl)-L-homocysteine, TLC monitoring the reactions for 6 to 10 hours, washing to remove impurities, and carrying out extraction, drying, and crystallization t obtain a solid namely N-fluorenylmethoxycarbonyl-S-(4-methoxytribenzyl)-L-homocysteine.
Poly(S-ethylsulfonyl- l -homocysteine): An α-Helical Polypeptide for Chemoselective Disulfide Formation
Muhl, Christian,Sch?fer, Olga,Bauer, Tobias,R?der, Hans-Joachim,Barz, Matthias
, p. 8188 - 8196 (2018/10/31)
Homocysteine and cysteine are the only natural occurring amino acids that are capable of disulfide bond formations in peptides and proteins. The chemoselective formation of asymmetric disulfide bonds, however, is chemically challenging and requires an activating group combining stability against hard nucleophiles, e.g., amines, with reactivity toward thiols and soft nucleophiles. In light of these considerations, we introduced the S-alkylsulfonyl cysteines in our previous work. Here, we present the synthesis and ring-opening polymerization of S-ethylsulfonyl-l-homocysteine N-carboxyanhydrides. We demonstrate that the polymerization leads to narrowly distributed polypeptides (D= 1.1-1.3) with no detectable side reactions in a chain length regime from 11 to 165. In contrast to the already reported cysteine derivatives, poly(S-ethylsulfonyl-l-homocysteine)s do not form β-sheets, which reduce solubility and limit the degree of polymerization of poly(S-ethylsulfonyl-l-cysteine)s to 50. Instead, these polymers form α-helices as confirmed by circular dicroism (CD) experiments and infrared spectroscopy (FT-IR). In comparison to the cysteine derivatives, the α-helix formation leads to slightly faster polymerization kinetics (rate constants from 1.44 × 10-5 to 5.29 × 10-5 s-1). In addition, the ability for the chemoselective formation of asymmetric disulfides is preserved as monitored via 1H NMR experiments. Consequently, this new polypeptide overcomes the chain length limitations of poly(S-ethylsulfonyl-l-cysteine)s and thus provides convenient access to reactive poly(S-ethylsulfonyl-l-homocysteine)s for chemoselective disulfide formation.
A role for glutamate-333 of Saccharomyces cerevisiae cystathionine γ-lyase as a determinant of specificity
Hopwood, Emily M.S.,Ahmed, Duale,Aitken, Susan M.
, p. 465 - 472 (2014/01/17)
Cystathionine γ-lyase (CGL) catalyzes the hydrolysis of l-cystathionine (l-Cth), producing l-cysteine (l-Cys), α-ketobutyrate and ammonia, in the second step of the reverse transsulfuration pathway, which converts l-homocysteine (l-Hcys) to l-Cys. Site-directed variants substituting residues E48 and E333 with alanine, aspartate and glutamine were characterized to probe the roles of these acidic residues, conserved in fungal and mammalian CGL sequences, in the active-site of CGL from Saccharomyces cerevisiae (yCGL). The pH optimum of variants containing the alanine or glutamine substitutions of E333 is increased by 0.4-1.2 pH units, likely due to repositioning of the cofactor and modification of the pKa of the pyridinium nitrogen. The pH profile of yCGL-E48A/E333A resembles that of Escherichia coli cystathionine β-lyase. The effect of substituting E48, E333 or both residues is the 1.3-3, 26-58 and 124-568-fold reduction, respectively, of the catalytic efficiency of l-Cth hydrolysis. The Kml-Cth of E333 substitution variants is increased ~ 17-fold, while Km l-OAS is within 2.5-fold of the wild-type enzyme, indicating that residue E333 interacts with the distal amine moiety of l-Cth, which is not present in the alternative substrate O-acetyl-l-serine. The catalytic efficiency of yCGL for α,γ-elimination of O-succinyl-l-homoserine (k cat/Kml-OSHS = 7 ± 2), which possesses a distal carboxylate, but lacks an amino group, is 300-fold lower than that of the physiological l-Cth substrate (kcat/Kml-Cth = 2100 ± 100) and 260-fold higher than that of l-Hcys (k cat/Kml-Hcys = 0.027 ± 0.005), which lacks both distal polar moieties. The results of this study suggest that the glutamate residue at position 333 is a determinant of specificity.
A green and expedient synthesis of enantiopure diketopiperazines via enzymatic resolution of unnatural amino acids
Pereira, Pedro C.,Arends, Isabel W.C.E.,Sheldon, Roger A.
supporting information, p. 4991 - 4993 (2015/01/09)
Dipeptides comprising a d-phenylglycyl moiety coupled to the l-enantiomer of 2-amino butyric acid, norvaline, norleucine, and homocysteine were successfully synthesized from d-phenylglycine amide and the racemate of the corresponding unnatural amino acid.
BINARY AND TERTIARY GALVANIC PARTICULATES AND METHODS OF MANUFACTURING AND USE THEREOF
-
, (2012/06/18)
The present invention relates to galvanic particulates, their methods of manufacture and uses in treatments are described. The galvanic particulates may be binary or tertiary galvanic particulates, for example, containing multiple layers or phases of conductive materials.
