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1.
《Biochimica et Biophysica Acta (BBA)/General Subjects》1986,882(2):258-265
In microsomal vesicles isolated from several smooth muscles many polypeptides were phosphorylated by the catalytic subunit of cyclic AMP-dependent protein kinase. In pig stomach and in rabbit and dog aorta components of Mr 22 000 and 11 000 were identified as forms of phospholamban. These polypeptides were, however, not observed in pig aorta. These phospholamban-like polypeptides presented the same electrophoretic mobility in sodium dodecyl sulphate gels as cardiac phospholamban, and the 22 000 Mr form showed a similar reaction to heat treatment in sodium dodecyl sulphate. Antibodies against purified canine cardiac phospholamban cross-reacted with the 22 000 and 11 000 Mr phosphorylatable polypeptides from smooth muscle membranes. Subcellular fractionation of porcine stomach smooth muslce indicated that phospholamban was present in the membranes of the endoplasmic reticulum and not in the plasma membranes. Phospholamban was also phosphorylated by an endogenous calcium—calmodulin-dependent protein kinase and by an endogenous cyclic AMP-dependent kinase. It is concluded that the endoplasmic reticulum of many, but possibly not all, smooth muscles contains phospholamban. However, the physiological role of phospholamban in smooth muscle remains to be established. 相似文献
2.
Phosphoglucose isomerase is the first committed enzyme of glycolysis. The protein also has a variety of biological activities on mammalian cells. The molecular basis of these extracellular functions is unclear, and the high resolution three-dimensional structure of a mammalian enzyme has not been described. We report here the cDNA and protein sequence for phosphoglucose isomerase from rabbit muscle. The sequence was obtained directly by PCR without the need to screen clones from a cDNA library and encoded active enzyme when expressed in bacterial cells. The 558 amino acid rabbit coding sequence is the same length as and highly similar (92% residue identity) to the sequences from human and pig and less so (88%) to the mouse enzyme. Non-conservative amino acid changes between the four mammalian sequences are concentrated in the first 35 and last five residues. The rabbit protein has an additional Cys residue and amino acid changes at five positions otherwise invariant in the mammalian enzymes. 相似文献
3.
4.
Expression of endoplasmic-reticulum Ca2(+)-pump isoforms and of phospholamban in pig smooth-muscle tissues. 总被引:3,自引:0,他引:3
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The expression of the gene 2 sarcoplasmic/endoplasmic-reticulum Ca2(+)-pump isoforms (SERCA2a and SERCA2b) and of phospholamban was studied in pig smooth muscle of the stomach, longitudinal ileum, pulmonary artery and aorta. mRNA levels were determined using an RNAase protection assay. The SERCA2 isoforms and phospholamban were tested on Western blots with a panel of antibodies, some of which were isoform-specific. The pig smooth-muscle tissues all contained comparable SERCA2 mRNA levels, but these levels were 10-20-fold lower than SERCA2 mRNA levels in cardiac muscle. Of the SERCA2 mRNAs in smooth muscle, 72-81% encoded the non-muscle isoform (SERCA2b), and Western blot analysis with isoform-specific antibodies confirmed that the SERCA2b isoform is the predominant endoplasmic-reticulum Ca2(+)-pump in smooth muscle. In contrast with SERCA2 mRNA levels, phospholamban mRNA levels varied by 12-fold between the different pig smooth-muscle tissues, with low and very low levels in the pig pulmonary artery and the pig aorta respectively. The differential expression of phospholamban was also confirmed on Western blots. The finding that the phospholamban content varied between the different smooth-muscle tissues whereas the SERCA2 expression remained rather constant indicates that, in pig smooth muscle, the expression of phospholamban is not coupled with that of SERCA2. 相似文献
5.
Summary. The nucleotide sequence of cDNA that encodes hamster d-amino-acid oxidase (DAO) was determined. The cDNA consisted of 1,590 nucleotides and a poly(A) tail. It had an open reading
frame for a protein consisting of 346 amino acid residues. The number of the amino acid residues is the same as that of the
rat DAO. However, the hamster DAO has one residue more than mouse DAO and one residue less than human, pig, rabbit, and guinea
pig DAOs. Amino acid sequence of the hamster DAO was highly similar to those of mouse and rat DAOs: 89% and 88% of the amino
acid residues were identical between the hamster and mouse DAOs and between the hamster and rat DAOs, respectively. The homology
was slightly less between the hamster DAO and the human (81%), pig (78%), rabbit (78%), or guinea pig DAO (82%). It has been
proposed that the mouse and rat DAOs lack an amino acid residue corresponding to the 25th residue of the DAOs of other mammals.
However, a detailed comparison of the amino acid sequences as well as the underlying nucleotide sequences by inclusion of
the hamster ones revealed that the rodent DAOs does not lack the 25th, but the 27th residue.
Received January 16, 2002 Accepted June 20, 2002 Published online November 14, 2002
Authors' address: Dr. Ryuichi Konno, Department of Microbiology, Dokkyo University School of Medicine, Mibu, Tochigi 321-0293, Japan, Fax:
+81-282-86-5616 相似文献
6.
Substrate determinants for rabbit and chicken skeletal muscle myosin light chain kinases were examined with synthetic peptides. Both skeletal muscle myosin light chain kinases had similar phosphorylation kinetics with synthetic peptide substrates. Average kinetic constants for skeletal muscle myosin light chain heptadecapeptide, (formula; see text) where S(P) is phosphoserine, were Km, 2.3 microM and Vmax, 0.9 mumol/min/mg of enzyme. Km values were 122 and 162 microM for skeletal muscle peptides containing A-A for basic residues at positions 2-3 and 6-7, respectively. Average kinetic constants for smooth muscle myosin light chain peptide, (formula; see text), were Km, 1.4 microM and Vmax 27 mumol/min/mg of enzyme. Average Km values for the smooth muscle peptide, residues 11-23, were 10 microM which increased 6- and 11-fold with substitutions of alanine at residues 12 and 13, respectively. Vmax values decreased and Km values increased markedly by substitution of residue 16 with glutamate in the 11-23 smooth muscle tridecapeptide. Basic residues located 3 and 6-7 residues toward the NH2 terminus from phosphoserine in smooth muscle myosin light chain and 6-8 and 10-11 residues toward the NH2 terminus from phosphoserine in skeletal muscle myosin light chain appear to be important substrate determinants for skeletal muscle myosin light chain kinases. These properties are different from myosin light chain kinase from smooth muscle. 相似文献
7.
Molecular cloning of cDNA encoding the Ca2+ release channel (ryanodine receptor) of rabbit cardiac muscle sarcoplasmic reticulum 总被引:28,自引:0,他引:28
K Otsu H F Willard V K Khanna F Zorzato N M Green D H MacLennan 《The Journal of biological chemistry》1990,265(23):13472-13483
We have cloned and sequenced cDNA encoding the Ca2+ release channel (ryanodine receptor) of rabbit cardiac muscle sarcoplasmic reticulum. The cDNA, 16,532 base pairs in length, encodes a protein of 4,969 amino acids with a Mr of 564,711. The deduced amino acid sequence is 66% identical with that of the skeletal muscle ryanodine receptor, but analysis of predicted secondary structures and hydropathy plots suggests that the two isoforms exhibit the same topology in both transmembrane and cytoplasmic domains. A potential ATP binding domain was identified at residues 2619-2652, a potential phosphorylation site at residue 2809, and potential calmodulin binding sites at residues 2775-2807, 2877-2898, and 2998-3016. We suggest that a modulator binding domain in the protein lies between residues 2619 and 3016. Northern blot analysis of mRNA from a variety of tissues demonstrated that the cardiac isoform is expressed in heart and brain, while the skeletal muscle isoform is expressed in both fast- and slow-twitch muscle. No ryanodine receptor mRNA was detected in extracts from smooth muscle or any other non-muscle tissue examined. The two receptors are clearly the products of separate genes, and the gene encoding the cardiac muscle ryanodine receptor was localized to chromosome 1. 相似文献
8.
Sequencing of the chicken non-erythroid spectrin cDNA reveals an internal repetitive structure homologous to the human erythrocyte spectrin. 总被引:5,自引:0,他引:5
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Immunological screening of a chicken gizzard cDNA expression library was used to isolate two clones encoding a part of the non-erythroid spectrin-like protein. Clones were identified by immunoblotting of the polypeptides synthesized in Escherichia coli cells transformed with cDNA cloned in the pUC8 plasmid vector using polyclonal rabbit antibodies raised against bovine non-erythroid spectrin. The sequence of an approximately 1.5-kb cDNA insert of one clone was determined. Analysis of the predicted amino acid sequence reveals that, despite differences in immunological cross-reactivity and peptide maps, the chicken non-erythroid and the human erythrocyte spectrins are highly homologous proteins. Like the human erythrocyte spectrin, the chicken smooth muscle spectrin appears also to be constructed from repeated, homologous structures of 106 amino acid residues. This is probably a universal structure motif of spectrins. 相似文献
9.
Sweet potato (Ipomoea batatas) starch phosphorylase cDNA clones were isolated by screening an expression library prepared from the young root poly(A)+ RNA successively with an antiserum, a monoclonal antibody, and a specific oligonucleotide probe. One cDNA clone had 3292 nucleotide residues in which was contained an open reading frame coding for 955 amino acids. This sequence was compared with those of potato (916 residues plus 50-residue putative transit peptide) and rabbit muscle (841 residues) phosphorylases. The sweet potato phosphorylase has an overall structural feature highly homologous to that reported for potato phosphorylase, in conformity with the finding that they belong to the same class of plant phosphorylase. High divergencies of the two enzymes are found in the about 70 residue N-termini each including a putative transit peptide, and the midchain 78 residue insert typical of type I plant phosphorylase. We consider that the very high dissimilarity found in the midchain inserts is related to the difference in proteolytic lability of the two plant phosphorylases. Some structural features of the cDNA clone were also discussed. 相似文献
10.
cDNA cloning of human calpastatin: sequence homology among human, pig, and rabbit calpastatins 总被引:4,自引:0,他引:4
K Asada Y Ishino M Shimada T Shimojo M Endo F Kimizuka I Kato M Maki M Hatanaka T Murachi 《Journal of enzyme inhibition》1989,3(1):49-56
cDNA of human calpastatin, an inhibitor protein specific for calpain (EC 3.4.22.17; Ca2(+)-dependent cysteine proteinase) was isolated by screening of a library prepared from human liver mRNA with pig calpastatin cDNA fragment as a probe. The primary structure of human calpastatin was deduced from the nucleotide sequence of the cDNA and compared with that of pig and rabbit calpastatins already reported. Human calpastatin consisted of 673 amino acid residues and had 78% and 77% identity to pig or rabbit calpastatins, respectively. Human calpastatin had a domain structure with four internally repetitive sequences and one N-terminal non-homologous sequence like the other calpastatins. Human calpastatin had two deletions, 22 and 13 residues long in domain L and domain 1, respectively, compared to pig or rabbit calpastatins. 相似文献
11.
Dog apolipoprotein (apo) C-I, A-I, and E cDNA clones were identified in a dog liver cDNA library in lambda gt10 by hybridization to synthetic oligonucleotide probes with the corresponding human DNA sequences. The longest clone for each apolipoprotein was completely sequenced. The apoC-I cDNA sequence predicts a protein of 62 residue mature peptide preceded by a 26 amino acid signal peptide. The apoA-I cDNA sequence predicts a 242 residue mature peptide, a 6 residue pro-segment, and an 18 residue signal peptide. The apoE cDNA, which lacks the signal peptide region, predicts a mature peptide of 291 amino acid residues. Slot blot hybridization of total RNA isolated from various dog tissues to dog apoC-I, A-I, and E cDNA probes indicates that apoC-I mRNA is detectable in liver only, apoA-I mRNA is present in liver and small intestine, though the concentration in the latter tissue is only approximately 15% of that in the liver, and apoE mRNA is present in multiple tissues including liver, jejunum, urinary bladder, ileum, colon, brain, kidney, spleen, pancreas, and testis with relative concentrations (%) of 100, 17.5, 7.5, 6.9, 5.9, 5.5, 5.0, 3.3, 1.0, and 1.0, respectively. These tissue distributions indicate that nascent lipoprotein particles produced in the dog small intestine would contain apoA-I and apoE but not apoC-I. The widespread tissue distribution of apoE mRNA indicates that like other mammals, peripheral synthesis of apoE contributes significantly to the total apoE pool in dog. We next compared the cDNA sequences among different vertebrate species for apoC-I (human and dog), A-I (human, rat, dog, rabbit and chicken), and E (human, rat, dog and rabbit) and calculated the rate of nucleotide substitution for each gene. Our results indicate that apoC-I has evolved rather rapidly and that on the whole, apoA-I is more conservative than apoE, contradictory to an earlier suggestion. ApoA-I is also more conservative than a region (residues 4204-4536) at the carboxyl-terminal portion, but less conservative than a region (residues 595-979) at the amino-terminal portion of apoB-100. Some regions in each of the apolipoproteins studied are better conserved than others and the rate of evolution of individual regions seems to be related to the stringency of functional requirements. Finally, we estimate that the human apoC-I pseudogene arose more than 35 million years ago, becoming nonfunctional soon after its formation. 相似文献
12.
Purification and characterization of Contractin A from the pedicellarial venom of sea urchin, Toxopneustes pileolus 总被引:1,自引:0,他引:1
A component that causes contraction of the isolated guinea pig tracheal smooth muscle was isolated in homogeneous form from the venom of the pedicellaria of the sea urchin, Toxopneustes pileolus. It is named Contractin A. Contractin A has 18,000 Da with a total residue of 138 amino acids. The molecular weight is about 17,700. The N-terminal amino acid is serine. The partial amino acid sequence was determined up to 37 residues. Direct comparison of sea urchin Contractin A does not show any similarity in amino acid sequence to toxins isolated from other marine toxin producers such as sea snakes, sea anemones, or marine worms. Contractin A caused contraction of the tracheal smooth muscle in a dose-dependent manner. Furthermore, Contractin A relaxed the contraction induced by histamine. The contraction and relaxation activity of Contractin A on the tracheal smooth muscle is reduced by a cyclooxygenase inhibitor such as indomethacin. The contraction induced by Contractin A is also inhibited by a phospholipase C inhibitor but not by a phospholipase A2 inhibitor. These results suggest that in the isolated guinea pig tracheal smooth muscle, the response to Contractin A may be effected through activated phospholipase C. 相似文献
13.
Apolipoprotein C-II (apoC-II) plays a critical role in the metabolism of plasma lipoproteins as an activator for lipoprotein lipase. Human apoC-II consists of 79 amino acid residues (pro-apoC-II). A minor fraction is converted to a mature form by cleavage at the site QQDE releasing the 6 amino-terminal residues. We have cloned and sequenced the cDNA for rat apoC-II from a liver cDNA library using human apoC-II cDNA as a probe. The cDNA encodes a protein of 97 amino acid residues including a signal peptide of 22 amino acid residues. There is approximately 60% similarity between the deduced amino acid sequence of rat apoC-II and other apoC-II sequences presently known (human, monkey, dog, cow, and guinea pig). Compared to these, rat apoC-II is one residue shorter at the carboxyl terminus. Furthermore, there is a deletion of 3 amino acid residues (PQQ) in the highly conserved cleavage site where processing from pro- to mature apoC-II occurs in other species. Accordingly, rat apoC-II isolated from plasma was mainly in the pro-form. Northern blot analyses indicated that rat apoC-II is expressed both in liver and in small intestine. 相似文献
14.
M.M.T. O''Hare S. Tenmoku L. Aakerlund L. Hilsted A. Johnsen T.W. Schwartz 《Regulatory peptides》1988,20(4):293-304
Neuropeptide Y (NPY) was isolated and characterised from acid-ethanol extracts of rabbit and guinea pig brain. In both instances the chromatographic purification was a two-step procedure of gel filtration followed by reverse-phase high-performance liquid chromatography. The amino acid sequence of rabbit and guinea pig NPY was found to be identical to human and rat NPY as deduced from the cDNA structures. With the exception of the porcine peptide, all mammalian NPYs characterised to date have a methionine residue in position 17. This methionine residue is readily oxidized as indicated by the high degree of spontaneous oxidation of peptides found in the rabbit and guinea pig brain extracts and in NPY extracted from a rat phaeochromocytoma cell line. It is concluded that NPY is among the most highly conserved peptides and that NPYs containing methionine in position 17 are prone to oxidation. 相似文献
15.
Suzuki O Mochida K Yamamoto Y Noguchi Y Takano K Matsuda J Ogura A 《Molecular reproduction and development》2002,62(3):335-342
The common alpha-subunit of glycoprotein hormones (CGalpha) is a core protein shared by follicle-stimulating hormone (FSH), luteinizing hormone (LH), and thyroid-stimulating hormone (TSH). In order to obtain a molecular basis for an efficient superovulation technique applicable to a wide range of animal species and to discuss the phylogenetic aspect based on molecules related to the reproductive system, we determined cDNA sequences of CGalpha in seven laboratory animals: the guinea pig, Mongolian gerbil, golden hamster, mastomys, Japanese field vole, the JF1 strain of Mus musculus molossinus, and rabbit. Comparison of the inferred CGalpha amino acid sequences of these animals and other mammals (human, mouse, rat, cow, pig, and sheep) showed that the signal peptides and the first ten residues at the N-terminus of the apoprotein were variable, while the rest of the apoproteins were highly conserved. In particular, all rodents had a leucine residue at the apoprotein N-terminus, except the guinea pig, which had a phenylalanine residue, as in the cow, pig, sheep, and rabbit. Phylogenetic trees constructed from amino acid sequences suggest a closer relationship between the guinea pig and artiodactyls than to rodents, confirming the taxonomic peculiarity of the guinea pig. 相似文献
16.
4-Aminobutyrate aminotransferase is a key enzyme of the 4-aminobutyric acid shunt. It is responsible for the conversion of the neurotransmitter 4-aminobutyrate to succinic semialdehyde. By using oligonucleotide probes based on partial amino acid sequence data for the pig brain enzyme, several overlapping cDNA clones of 2.0-2.2 kilobases in length have been isolated. The largest cDNA clone was selected for sequence analysis. The amino acid sequence predicted from the cDNA sequence shows that the precursor of 4-aminobutyrate aminotransferase consists of the mature enzyme of 473 amino acid residues and an amino-terminal segment of 27 amino acids attributed to the signal peptide. The cofactor pyridoxal-5-P is bound to lysine residue 330 of the deduced amino acid sequence of the mature enzyme. 相似文献
17.
Complementary DNA clones specific for phospholamban have been isolated from a canine cardiac cDNA library. The amino acid sequence deduced from the cDNA sequence showed that phospholamban consisted of 52 amino acid residues and was synthesized without an amino-terminal signal sequence. The RNA blot analysis revealed that phospholamban mRNAs were represented by two main species of approximately 1.2kb and approximately 2.8kb. These mRNAs appeared to differ primarily in the length of the 3' untranslated region. 相似文献
18.
Substrate specificity of myosin light chain kinases. 总被引:3,自引:0,他引:3
Skeletal muscle myosin light chain kinase can phosphorylate myosin light chains isolated from skeletal or smooth muscle. In contrast, smooth muscle myosin light chain kinase specifically phosphorylates light chains isolated from smooth muscle. In this study, we have identified residues within the rabbit smooth and skeletal muscle myosin light chain kinases which may interact with the basic residues that are important substrate determinants in the light chains. Mutation of aspartic acid 270 amino-terminal of the catalytic core of the skeletal muscle myosin light chain kinase increased the Km value for both smooth and skeletal muscle light chains. Although deletions of the analogous region of the smooth muscle myosin light chain kinase (residues 663-678) markedly increased the Km value for light chain, mutation of any single acidic residue within this region did not have a similar effect. Mutation of single residues within the catalytic core of the skeletal muscle (E377 and E421) and smooth muscle (E777 and E821) myosin light chain kinases increased Km values for the smooth muscle light chain at least 35- and 100-fold, respectively. It is proposed that these residues may form ionic interactions with the arginine that is 3 residues amino-terminal of the phosphorylatable serine in the smooth muscle light chain. 相似文献
19.
P T Cohen 《FEBS letters》1988,232(1):17-23
20.
Biological activities of bovine cardiac-muscle troponin C C-terminal peptide (residues 84-161).
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1. Bovine cardiac-muscle troponin C was digested at cysteine residues 35 and 84, and the C-terminal peptide (residues 84-161) was isolated. 2. The C-terminal peptide contains two Ca2+-binding sites. These sites bind Ca2+ with a binding constant of 2.0 X 10(8) M-1. In the presence of 2 mM-Mg2+ the binding constant for Ca2+ is decreased to 3.7 X 10(7) M-1. The corresponding constants for native troponin C are 5.9 X 10(7) M-1. and 2.9 X 10(7) M-1 respectively. 3. Electrophoretic mobility of the C-terminal peptide is increased in the presence of 0.1 mM-CaCl2 as compared with the mobility in the presence of 2mM-EDTA. The same phenomenon was observed when electrophoresis was performed in the presence of 6 M-urea or 0.1% sodium dodecyl sulphate. 4. When saturated with Ca2+, the C-terminal peptide forms complexes with bovine cardiac-muscle troponin I both in the absence and in the presence of 6 M-urea. This complex is dissociated on removal of Ca2+. 5. The data suggest that the C-terminal peptide of troponin C contains two Ca2+/Mg2+-binding sites and interacts with troponin I. Thus, despite the 30% difference in amino acid composition, the properties of bovine cardiac-muscle troponin C C-terminal peptide are similar to those of rabbit skeletal-muscle troponin C C-terminal peptide. 相似文献