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1.
Rat ovarian angiotensin II receptors. Characterization and coupling to estrogen secretion 总被引:6,自引:0,他引:6
Angiotensin II receptor agonist (125I-angiotensin II) and antagonist (125I-[Sar1,Ile8]angiotensin II) bind in a specific and saturable manner to rat ovarian membranes. Agonist and antagonist binding affinity (KD approximately 0.5 nM) and the number of sites estimated (Bmax approximately 60 fmol/mg of protein) were similar. Dissociation of receptor-bound agonist was more rapid than the dissociation of receptor-bound antagonist, and agonist, but not antagonist, dissociation from the receptor was accelerated by GTP gamma S. A 0-150 mM increase in Na+ produced a 27% increase in the KD of agonist binding. Antagonist binding was not modified by Na+. These studies suggest that both agonist and antagonist identify putative angiotensin II receptors in the ovary but that the properties of agonist and antagonist binding are distinct. Angiotensin II antagonist binding sites are present on the granulosa cell layer of rat ovarian follicles (Speth, R. C., Bumpus, F. M., and Husain, A. (1986) Eur. J. Pharmacol. 130, 351-352). To determine the role of angiotensin II in ovarian function, we examined angiotensin II receptors and function during the onset of puberty. High affinity and low capacity angiotensin II receptors were present in ovaries from immature rats. After pregnant mare's serum gonadotropin induced ovulation in immature rats, antagonist binding to total ovarian membranes increased over 3-fold. In vitro incubation of peripubertal ovaries with 1 microM angiotensin II produced a stimulation of estrogen, but not progesterone, secretion. This steroidogenic effect of angiotensin II was most pronounced in the luteal phase of the estrus cycle. These studies point toward the involvement of angiotensin II in the regulation of ovarian function, possibly through modulation of follicular estrogen levels. 相似文献
2.
P A Husain J E Colbert S R Sirimanne D G VanDerveer H H Herman S W May 《Analytical biochemistry》1989,178(1):177-183
A chiral derivatizing reagent, N-succinimidyl-2-(S)-methoxy-2-phenylacetic acid ester (SMPA), directed toward reaction with primary amine-containing compounds has been synthesized and characterized. This reagent is suitable for HPLC resolution from enzymatic-scale reactions where only microgram quantities of chiral products may be obtainable. SMPA derivatization was shown to be effective in the resolution of the enantiomers of a number of different racemic compounds. SMPA was used to resolve the diastereoisomeric derivatives of a previously unknown enzymatically oxygenated product, allowing determination of the stereochemical course of the enzymatic reaction. SMPA is easily prepared from an inexpensive, commercially available, and enantiomerically pure precursor with the formation of a shelf-stable crystalline product which is utilizable in water-containing solutions. In addition to its usefulness for micro-determinations, SMPA is useful for preparative-scale resolutions of enantiomers since the reagent is cleaved from the diastereoisomeric derivative by acid hydrolysis. 相似文献
3.
Unusual redox properties of electron-transfer flavoprotein from Methylophilus methylotrophus 总被引:1,自引:0,他引:1
The most positive redox potential ever recorded for a flavin adenine dinucleotide (FAD) containing protein has been measured for an electron-transfer flavoprotein (ETF) synthesized by Methylophilus methylotrophus. This potential value, 0.196 V versus the standard hydrogen electrode (vs SHE), was measured at pH 7.0 for the one-electron reduction of fully oxidized ETF (ETFox) to the red anionic semiquinone form of ETF (ETF.-). Quantitative formation of ETF.- was observed. The first successful reduction of ETF from M. methylotrophus to its two-electron fully reduced form was also achieved. Although addition of the second electron to ETF.- was extremely slow, the potential value measured for this reduction was -0.197 V vs SHE, suggesting a kinetic rather than thermodynamic barrier to two-electron reduction. These data are believed to be consistent with the postulated catalytic function of ETF to accept one electron from the iron-sulfur cluster of trimethylamine dehydrogenase (TMADH). The second electron reduction appears to have no catalytic function. The very positive potential measured for this ETF and the wide separation of potentials for the two electron reduction steps show that this ETF is a unique and interesting flavoprotein. In addition, this work highlights that while ETFs exhibit similar structural and spectral properties, they display wide variations in redox properties. 相似文献
4.
A thermotolerant methylotrophicBacillus sp. (KISRI TM1A, NCIMB 40040), isolated from the Kuwaiti environment and belonging to the group II spore-forming, bacilli, could not be correlated with any knownBacillus sp. It may, therefore, be a new species. It grew at temperatures from 37° to 58°C from pH 6.5 to 9.0 and on methanol up to 40 g l–1. It grew well in a chemostat. Its biomass yield coefficient was improved by about 30% by optimization of medium and growth conditions, reaching a maximum of 0.44g g–1 at 45°C pH 6.8 to 7.0, dilution rate 0.25 h–1 with methanol at 10 g l–1. Average crude protein and amino acid content were 84% and 60%, respectively, and maximum productivity attained under laboratory conditions was 5.06 g l–1h–1. It was concluded that this strain has good potential for use in single-cell protein production. 相似文献
5.
K D Sharma T M Loehr J Sanders-Loehr M Husain V L Davidson 《The Journal of biological chemistry》1988,263(7):3303-3306
The copper binding site of amicyanin from Paracoccus denitrificans has been examined by resonance Raman spectroscopy. The pattern of vibrational modes is clearly similar to those of the blue copper proteins azurin and plastocyanin. Intense resonance-enhanced peaks are observed at 377, 392, and 430 cm-1 as well as weaker overtones and combination bands in the high frequency region. Most of the peaks below 500 cm-1 shift 0.5-1.5 cm-1 to lower energy when the protein is exposed to D2O. Based on the pattern of conserved amino acids, the axial type EPR spectrum, and the resonance Raman spectrum, it is proposed that the copper binding site in amicyanin contains a Cu(II) ion in a distorted trigonal planar geometry with one cysteine and two histidine ligands and an axial methionine ligand at a considerably longer distance. Furthermore, the presence of multiple intense Raman peaks in the 400 cm-1 region which are sensitive to deuterium substitution leads to the conclusion that the Cu-S stretch is coupled with internal ligand vibrational modes and that the sulfur of the cysteine ligand is likely to be hydrogen-bonded to the polypeptide backbone. 相似文献
6.
Properties of Paracoccus denitrificans amicyanin 总被引:4,自引:0,他引:4
Paracoccus denitrificans synthesizes an inducible, periplasmic, blue copper protein [Husain, M., & Davidson, V.L. (1985) J. Biol. Chem. 260, 14626-14629] that can be classified as an amicyanin on the basis of its ability to accept electrons from methylamine dehydrogenase. The amino acid composition and sequence of the 10 N-terminal residues of this protein have been determined. From these data, it is evident that amicyanin is structurally distinct from azurins as it contains no disulfide bond and an N-terminal sequence that is completely different from the highly conserved N-terminal azurin sequences. Dialysis of reduced amicyanin against potassium cyanide resulted in a nearly quantitative yield of apoamicyanin. Amicyanin and apoamicyanin exhibit fluorescence emission maxima at 314 nm when excited at 280 nm. Addition of 6 M guanidine hydrochloride shifts these emission maxima to 350 nm. The fluorescence intensity of apoamicyanin is 10-fold greater than that of amicyanin. Addition of copper to the apoprotein caused a stoichiometric quenching of fluorescence and restoration of visible absorbance with no concomitant change in absorbance at 280 nm. At least one cysteine residue, which reacts with 5,5'-dithiobis(2-nitrobenzoic acid) in apoamicyanin, does not react in the holoprotein, even in the presence of 6 M guanidine hydrochloride. Reductive and oxidative titrations of amicyanin indicate that it is a one-electron carrier. This amicyanin is also able to accept electrons from the methylamine dehydrogenase isolated from bacterium W3A1, which is taxonomically very different from P. denitrificans. 相似文献
7.
8.
Completion of the amino acid sequence of papain 总被引:8,自引:6,他引:2
Papain was inhibited with bromo[2-(14)C]acetic acid, the tertiary structure of the inhibited enzyme was unfolded and the disulphide bridges were reduced with mercaptoethanol and aminoethylated. Digestion with trypsin gave a radioactive peptide consisting of residues 18-58 inclusive and containing therefore the sequence of the thirteen unknown residues 29-41 in the primary sequence of papain. This peptide was digested with pepsin to give a radioactive peptide consisting of residues 18-47, which after digestion with 0.4m-hydrochloric acid gave a radioactive peptide consisting of residues 24-43 inclusive. Further digestion with 6m-hydrochloric acid gave peptides that were used to determine the sequence: Ser-Ala-Val-Val-Thr-Ile-Glx-Gly-Ile-Ile-Lys-Ile-Arg for the residues 29-41, so completing the amino acid sequence of papain. 相似文献
9.
Papain was irreversibly inhibited by 1,3-dibromoacetone, a reagent designed to react first with the active-site cysteine residue and subsequently with a second nucleophile. The molecular weight of the inhibited enzyme was indistinguishable from that of papain itself, and no evidence of dimeric or oligomeric species was found. The optical-rotatory-dispersion curves of chloroacetone-inhibited papain and 1,3-dibromoacetone-inhibited papain were essentially similar. Amino acid analysis of the 1,3-dibromo[2-14C]acetone-inhibited enzyme and the performic acid-oxidized material clearly showed that a cysteine and histidine residue had been alkylated through the thiol and N-1 of the imidazole group respectively. These groups must therefore be within 5å of each other in the tertiary structure of papain. Possible mechanistic implications are briefly discussed. 相似文献
10.
Ersilia Cassano Marco Macchia Mahmud Hamdan Paolo Rovero 《Letters in Peptide Science》1996,3(3):117-120
Summary We describe a new solid-phase strategy for the selective reduction of the C=N bond in peptide oximes using a trialkylsilane in trifluoroacetic acid. The reduction is performed directly on the resin-bound peptide, with concomitant cleavage of the peptide from the resin and deblocking of protected side chains. 相似文献