Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: EC:3.4.21.1 (chymotrypsin)
10,938 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Cytochrome c1 is a subunit of ubiquinol--cytochrome c reductase (EC 1.10.2.2). In Neurospora crassa wild type 74A grown in the presence of chloramphenicol, the subunit is inserted only into the bilayer of the mitochondrial inner membranes without associating with other proteins. From these modified membranes a monodisperse (cytochrome c1)-Triton complex was isolated by subjecting the Triton-solubilized membranes to affinity chromatography on immobilized cytochrome c. A water-soluble pentamer of cytochrome c1 was prepared from the (cytochrome c1)-Triton complex by removing the detergent. By limited proteolytic digestion of the cytochrome c1-Triton complex with chymotrypsin, a water-soluble monomeric cytochrome c1 was prepared which has a molecular weight of only 24 000 as compared to 31 000 of the membrane-bound cytochrome c1. The 24 000-Mr cytochrome c1 and the 31 000-Mr cytochrome c1 have same light absorption spectra and cytochrome-c-binding properties. These results are used to propose the following model. Cytochrome c1 consists of a large hydrophilic part and a small hydrophobic part. The hydrophilic part extends from the mitochondrial inner membrane into the intermembrane space. This part carries the heme and interacts with cytochrome c. The hydrophobic part anchors the cytochrome c1 to the bilayer.
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PMID:Membrane-bound and water-soluble cytochrome c1 from Neurospora mitochondria. 626 10

The complete primary structure of bovine heart cytochrome c1 was established by analyses of peptide fragments prepared by digestion using trypsin, staphylococcal protease, and chymotrypsin and by cyanogen bromide cleavage of cytochrome c1 and its derivatives. The total number of amino acid residues is 241, giving a molecular weight of 27,924 including the heme group. The NH2- and COOH-terminal residues are serine and lysine, respectively. One characteristic of the protein is that cytochrome c1 contains 43.6% hydrophobic residues and the polarity is estimated to be 41.1%. No clear homology was found between cytochrome c1 and other membranous proteins such as cytochrome b5 or the subunits of cytochrome oxidase for which sequences have been reported. Cytochrome c1 is predicted to have a high content of alpha-helix (46%). Partial sequence studies were also carried out on cytochrome c1 preparations obtained by different procedures and showed that there is no difference among the sequences of various preparations of cytochrome c1. The presence of a hydrophobic cluster near the COOH-terminal region indicates that the COOH-terminal region of cytochrome C1 associates with, or is buried in, the phospholipid bilayer of the mitochondrial membrane.
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PMID:Structural studies of bovine heart cytochrome c1. 628 15

The amino acid sequence of the soluble monohaem cytochrome c-556 from Agrobacterium tumefaciens, strain B2a, has been determined. The sequence was derived from peptides obtained by digestion of the apoprotein with trypsin and chymotrypsin, and by subdigestion of some of the peptides with Staphylococcus aureus protease and thermolysin. Sequencing of the various peptides was achieved by a combination of manual dansyl-Edman degradation and automatic liquid-phase sequence analysis. The main characteristic of this cytochrome is that the haem-binding sequence Cys-Xaa-Yaa-Cys-His occurs in the C-terminal region of the polypeptide chain, the first cysteine being located 11 residues ahead of the C-terminal lysine-122. As such, the protein belongs to cytochrome c sequence class II (sensu Ambler). The cytochrome c-556 is the first example known of a class II cytochrome of the low-spin type isolated from an obligate aerobic organism.
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PMID:The complete amino-acid sequence of the low-spin class II cytochrome c-556 from Agrobacterium tumefaciens strain B2a. 629 89

The orientation of the different subunits of complex III in the yeast inner mitochondrial membrane has been investigated by several different approaches. Immunoinhibition studies of cytochrome c reductase activity in intact mitoplasts and submitochondrial particles using IgG obtained from specific antisera against complex III, the iron-sulfur protein, core protein I, and core protein II suggested a transmembranous orientation of the complex with the antigenic sites of the iron-sulfur protein exposed on the cytoplasmic surface of the membrane. A lack of immunoinhibition was observed with the IgG against either core protein suggesting that these proteins may not be involved in catalysis. Digestion of mitoplasts with chymotrypsin indicated that the protein mass of cytochromes b and c1 protrudes from the cytoplasmic surface of the membrane; however, the hemes of cytochrome b appear to be buried within the membrane while the heme of cytochrome c1 is partially exposed on the chymotrypsin-sensitive portion of the polypeptide. By contrast, the iron-sulfur protein does not protrude from the membrane as it is completely resistant to chymotrypsin digestion. Labeling with the hydrophilic membrane-impermeant probe diazobenzenesulfonate suggests that core protein II is exposed on both sides of the membrane but protrudes into the matrix; while core protein I is within the membrane. Immunoprecipitation studies of sodium dodecyl sulfate and Triton X-100-solubilized mitochondria with subunit-specific antisera suggest that cytochromes b and c1 and core protein I are tightly associated in complex III. By contrast, the iron-sulfur protein and core protein II are loosely associated with the other subunits of the complex such that they are dissociated by low concentrations of detergent.
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PMID:Topographical orientation of complex III in the yeast mitochondrial membrane. 631 51

Reverse-phase high-pressure liquid chromatography has been used for the purification of some large cyanogen bromide peptides from flavocytochrome b2 fragment alpha. Acetonitrile gradients at acid and/or neutral pH using mu Bondapak C18 columns were useful for the smaller peptides (43 and 67 residues). The two larger ones, alpha CB1 and alpha CB2, could only be separated from each other by trifluoroacetic acid/1-propanol gradients on mu Bondapak-CN columns. The various systems tested are presented and compared. The elucidation of the amino acid sequence of alpha CB2 (95 residues), alpha CB3 (67 residues) and alpha CB4 (43 residues) is described. The fragments were digested with trypsin, chymotrypsin and Staphylococcus aureus V8 protease as necessary. Fragment alpha CB2 was also cleaved at the unique tryptophanyl bond with cyanogen bromide. Peptides were fractionated by Sephadex chromatography, thin-layer finger-printing and/or high-pressure liquid chromatography. Peptides were sequenced mostly in the liquid phase sequenator. The cyanogen bromide peptides could be ordered using information obtained previously, as well as additional data obtained in this work. Together with the previous elucidation of cytochrome b2 core sequence and of the hinge region [Guiard, B. and Lederer, F. (1976) Biochimie (Paris) 58, 305--316; Ghrir, R. and Lederer, F. (1981) Eur. J. Biochem. 120, 279--287], the present results enable us to present the complete sequence of fragment alpha (314 residues) with only three overlaps missing between cyanogen bromide peptides. Sequence comparisons with other known flavoproteins do not indicate any noticeable similarity. Structural predictions indicate an alteration of alpha helices and beta structure. The possibility that the non-heme-binding portion of fragment alpha could constitute a flavin-binding domain is discussed.
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PMID:Primary structure of flavocytochrome b2 from baker's yeast. Purification by reverse-phase high-pressure liquid chromatography and sequencing of fragment alpha cyanogen bromide peptides. 636 48

Extracts of whole porcine ovary stimulate BALB/3T3 [3H] DNA synthesis in BALB/3T3 cells. Biochemical characterization indicates the growth factor is associated with a soluble cationic protein that is labile to heat, stable to dithiothreitol treatment and elutes from Sephadex G-100 between chymotrypsin (25,000 Mr) and cytochrome C (12,700 Mr). Extracts of thecal tissue have growth-promoting activity in both granulosa and 3T3 cells. Conditioned medium from thecal but not granulosa cell cultures stimulates [3H] DNA synthesis in quiescent 3T3 and granulosa cells. The results indicate the presence of an endogenous protein ovarian growth factor found in the theca that stimulates granulosa cell proliferation in vitro.
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PMID:An endogenous ovarian growth factor which stimulates BALB/3T3 and granulosa cell proliferation. 665 80

In the present studies, a novel form of highly purified cytochrome P-450 (cytochrome P-452) isolated from the hepatic microsomes of clofibrate-pretreated rats has been compared to the major isozymes isolated from the hepatic microsomes of rats pretreated with phenobarbital (cytochrome P-450) and 2-naphthoflavone (cytochrome P-447) using a number of biochemical criteria. The results show that these three isozymes exhibit marked structural differences from each other as judged by a complete lack of immunochemical cross-reactivity between the isozymes and the heterologous rabbit serum antibodies using Ouchterlony double diffusion, and non-identity between the limited proteolytic digestion maps of the three isozymes obtained in the presence of chymotrypsin, papain and Staphylococcus aureus V8 proteases. Furthermore, the three isozymes exhibited clear differences in their monomeric molecular weights determined on calibrated sodium dodecyl sulphate/polyacrylamide gel electrophoresis in gels of varying acrylamide concentration. Substantial differences were also observed in the substrate specificities of the isozymes, which were reflected in differences in the turnover rates and positional selectivities of the hemoproteins for some model substrates. In addition, the isozymes differed in their substrate binding affinities and their ability to interact with purified hepatic microsomal cytochrome b5, as judged using difference spectrophotometry. Finally, subtle differences were detected in the ultraviolet visible absorbance spectra of the hemoproteins in the ferric, ferrous, and carbonmonoxyferrous states. Taken collectively, the above data provides compelling evidence that fundamental differences exist between these cytochrome P-450 isozymes, further establishing the uniqueness of the major form of cytochrome P-450 induced by clofibrate pretreatment.
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PMID:Multiple forms of hepatic cytochrome P-450. Purification, characterisation and comparison of a novel clofibrate-induced isozyme with other major forms of cytochrome P-450. 669 12

Chromatography of electrophoretically homogeneous cytochrome P-450LM4 from cholestyramine-treated rabbits on octylamine-Sepharose resulted in the isolation of two subfractions, cytochrome P-450LM4 I and cytochrome P-450LM4 II, with different catalytic properties. The original cytochrome P-450LM4 fraction catalyzed 7 alpha-hydroxylation of cholesterol, 12 alpha-hydroxylation of 5 beta-cholestane-3 alpha, 7 alpha-diol, 25-hydroxylation of 5 beta-cholestane-3 alpha, 7 alpha-diol and 5 beta-cholestane-3 alpha, 7 alpha, 12 alpha-triol, 6 beta-hydroxylation of testosterone, and demethylation of ethylmorphine. Cytochrome P-450LM4 I was inactive in cholesterol 7 alpha-hydroxylation, but catalyzed the other hydroxylations. Cytochrome P-450LM4 II catalyzed efficient cholesterol 7 alpha-hydroxylation. It also catalyzed the other hydroxylations, although at lower rates than cytochrome P-450LM4 I. Emulgen inhibited all steroid hydroxylase activities in cytochrome P-450LM4 II except the cholesterol 7 alpha-hydroxylase activity. Cytochrome P-450LM4 I and cytochrome P-450LM4 II showed the same apparent molecular weight and spectral properties as the original cytochrome P-450LM4 fraction. The two subfractions differed in amino acid composition. They produced similar but not identical one-dimensional peptide maps upon limited proteolysis with papain, chymotrypsin, and trypsin. The results show that cytochrome P-450LM4 from cholestyramine-treated rabbits contains at least two species with different amino acid compositions and different substrate specificities toward C27-steroids involved in biosynthesis of bile acids.
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PMID:Hydroxylations in biosynthesis of bile acids. Isolation of subfractions with different substrate specificity from cytochrome P-450LM4. 681 87

Flavocytochrome b2 from baker's yeast is a bifunctional tetrameric protein which carries two prosthetic groups, FMN and heme, per subunit of Mr 58 000. The amino terminus of the subunit is wrapped around the heme and constitutes the so-called cytochrome b2 core (Mr 11 000), homologous to cytochrome b5. It has been shown in the past that a number of proteases (yeast proteases, chymotrypsin) preferentially cleave the peptide chain at a point situated much further down the polypeptide chain than the C terminus of the heme-binding domain. Some enzymatic parameters are concomitantly modified, but not the quaternary structure. This paper describes the conditions for selective proteolysis of intact flavocytochrome b2 and of its various previously studied stable nicked forms by the protease from Staphylococcus aureus V8. Successive attack by a combination of two proteases is also described. We have established the amino acid sequence of the area where proteolytic attack takes places, and shown that chymotrypsin and S. aureus protease open only one bond, whereas yeast proteases remove five residues from the central part. The various nicked forms, some of which have lost up to 16 amino acid residues, have been enzymatically characterized. These and previous results lend support to, but do not prove, the idea that the flavodehydrogenase part of flavocytochrome b2 may be composed of two domains, linked by the region accessible to proteases. That area might constitute a hinge or rather a clasp between the domains.
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PMID:Study of a zone highly sensitive to proteases in flavocytochrome b2 from Saccharomyces cerevisiae. 703 12

A new isozyme of cytochrome P-450 has been purified to electrophoretic homogeneity from hepatic microsomes of rabbits treated chronically with ethanol. Several criteria indicate that the ethanol-inducible cytochrome, which has a minimal molecular weight of 51,000 and is designated form 3a on the basis of its relative electrophoretic mobility, is distinct from the known isozymes of P-450. As judged spectrally, the new isozyme is high spin in the oxidized state, as is form 4, but differs in that the spin state is unperturbed by nonionic detergents. The absolute spectrum of the ferrous carbonyl complex of form 3a is red shifted as compared to that of forms 2, 3b, 3c, 4, and 6 and exhibits a maximum at 452 nm. The amino acid composition of form 3a is different from that of the other isozymes, and both the NH2- and COOH-terminal sequences are distinct; form 3a has an NH2-terminal alanine and a carboxyl-terminal leucine residue. Peptide mapping by sodium dodecyl sulfate-polyacrylamide gel electrophoresis following treatment with papain, chymotrypsin, or Staphylococcus aureus V8 protease and by high performance liquid chromatography following trypsinolysis indicates that form 3a is a unique gene product. This cytochrome displays the highest activity of all of the rabbit isozymes in the oxidation of ethanol to acetaldehyde and the p-hydroxylation of aniline when reconstituted with NADPH-cytochrome P-450 reductase and phospholipid in the presence of NADPH and oxygen.
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PMID:Purification and characterization of a unique isozyme of cytochrome P-450 from liver microsomes of ethanol-treated rabbits. 708 77


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