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M Kamo

Publications and source records attributed to M Kamo.

At least 37 records · Page 2Linked to original sources
PubMed1997

Identification of domains on the 43 kDa chlorophyll-carrying protein (CP43) that are shielded from tryptic attack by binding of the extrinsic 33 kDa protein with photosystem II complex.

The structural association of the spinach 33 kDa extrinsic protein with the 43 kDa chlorophyll-carrying protein (CP43) in oxygen-evolving photosystem II (PS II) complexes was investigated by comparing the peptide mappings and N-terminal sequences of the trypsin-digested products of NaCl-washed PS II membranes, which bind the 33 kDa protein, with those of CaCl2-washed PS II membranes, which lack the 33 kDa protein. (1) Peptide from N-terminus to Arg26 of CP43, which is exposed to stromal side, was digested in both PS II membranes, independent of binding of the 33 kDa protein. (2) Peptide bond of Arg357-Phe358 located in the large extrinsic loop E of CP43, which is exposed to lumenal side, was cleaved by trypsin in CaCl2-washed PS II membranes but not in NaCl-washed PS II membranes. This indicates that the region around Arg357-Phe358 in loop E of CP43 is shielded from tryptic attack by binding of the 33 kDa protein to PS II. (3) Trypsin treatment of CaCl2-washed PS II membranes also cleaved peptide bond between Lys457 and Gly458 in C-terminal region of CP43, while no cleavage of this region was detected by trypsin treatment of NaCl-washed PS II membranes. This implies that a conformational change of the C-terminal region of CP43 which is exposed to stromal side occurred upon removal of the 33 kDa protein, which makes the C-terminal region accessible to trypsin. (4) Release of peptide from Gln60 to C-terminus of the alpha-subunit of cytochrome b-559 was detected only in trypsin treatment of CaCl2-washed PS II membranes, indicating that the C-terminal region of this subunit is shielded from tryptic attack by binding of the 33 kDa protein. (5) The PS II membranes, in which Arg357-Phe358, Lys457-Gly458 of CP43 and the C-terminal part of the cytochrome b-559 alpha-subunit had been cleaved by trypsin, was no longer able to bind the 33 kDa protein. This strongly suggests that a domain in loop E of CP43 and/or the C-terminal region of the cytochrome b-559 alpha-subunit are necessary for binding of the extrinsic 33 kDa protein to PS II.

Amino Acid Sequence↗
PubMed1997

Identification of domains on the extrinsic 33-kDa protein possibly involved in electrostatic interaction with photosystem II complex by means of chemical modification.

The extrinsic 33-kDa protein of photosystem II (PSII) was modified with various reagents, and the resulting proteins were checked for the ability to rebind to PSII and to reactivate oxygen evolution. While modification of more than eight carboxyl groups of aspartyl and glutamyl residues with glycine methyl ester did not affect the rebinding and reactivating capabilities, modification of amino groups of lysyl residues with either N-succinimidyl propionate or 2, 4,6-trinitrobenzene sulfonic acid or modification of guanidino groups of arginyl residues with 2,3-butanedione resulted in a loss of rebinding and reactivating capabilities of the 33-kDa protein. Moreover, the number of lysyl and arginyl residues susceptible to modification was significantly decreased when the protein was bound to PSII as compared with when it was free in solution, whereas the number of carboxyl groups modified was little affected. These results suggested that positive charges are important for the electrostatic interaction between the extrinsic 33-kDa protein and PSII intrinsic proteins, whereas negative charges on the protein do not contribute to such interaction. By a combination of protease digestion and mass spectroscopic analysis, the domains of lysyl residues accessible to N-succinimidyl propionate or 2,4, 6-trinitrobenzene sulfonic acid modification only when the 33-kDa protein is free in solution were determined to be Lys4, Lys20, Lys66-Lys76, Lys101, Lys105, Lys130, Lys159, Lys186, and Lys230-Lys236. These domains include those previously reported accessible to N-hydroxysuccinimidobiotin only in solution (Frankel and Bricker (1995) Biochemistry 34, 7492-7497), and may be important for the interaction of the 33-kDa protein with PSII intrinsic proteins.

Amino Acid Sequence↗
PubMed1997

Bond-specific chemical cleavages of peptides and proteins with perfluoric acid vapors: novel peptide bond cleavages of glycyl-threonine, the amino side of serine residues and the carboxyl side of aspartic acid residues.

Peptide bond cleavages by vapors composed of various from aqueous solutions of perfluoric acid were studied using synthetic peptides and proteins, and specific conditions were established for peptide bond cleavages including a novel cleavage of the glycyl-threonine bond. The peptide bonds on the aminosides of serine residues were cleaved by exposure to a vapor of 75% aqueous heptafluorobutyric acid at 30 or 50 degrees C for 24 h. Glycyl-threonine peptide bonds were cleaved with vapors of various concentrations (5, 75, and 90%) of heptafluorobutyric acid at 30-40 degrees C for 24 h. The peptide bonds on the carboxylsides of aspartic acid residues were cleaved by exposure to a vapor of 0.2% heptafluorobutyric acid at 90 degrees C for 4 to 24 h. The same vapor cleaved aspartyl-proline bonds under milder conditions such as at 60 degrees C for 16 h, under which the other aspartyl bonds were uncleaved. These specific chemical cleavages were applied to several proteins including newly characterized proteins.

Amino Acid Sequence↗
PubMed1997

In vitro skin penetration and degradation of enkephalin, elcatonin and insulin.

The work described in this paper was designed to evaluate the relevance of in vitro skin penetration studies of peptides across rat skin. The apparent penetration of three peptides, enkephalin, elcatonin and insulin, in the presence of enhancers was not seen in the in vitro method using Franz diffusion cells. However, when a protease inhibitor was mixed in the receptor fluid, the penetration of enkephalin and insulin was observed. Although insulin penetrated in the presence of enhancers, the penetration was extremely small in quantity and the cumulative amount did not increase with time. When the degradation of peptides in the receptor fluid of Franz cell was estimated, these peptides, especially enkephalin and insulin, were rapidly hydrolyzed and were almost completely lost within 3 h in the absence of an inhibitor, while elcatonin was slowly degraded. The addition of protease inhibitors, such as gabexate (20 mM), camostat (20 mM) or bile salt (taurocholate and deoxycholate, 10 mM), to the receptor fluid inhibited the degradation to a considerable extent, with the first-order rate constants decreased to one-tenth compared with the constants without inhibitors. From the inhibitory study using specific inhibitors, it was clarified that enkephalin and elcatonin were mainly hydrolyzed by aminopeptidases, endopeptidases and serine proteases in the viable skin. Consequently, the results obtained from the in vitro penetration studies without inhibitors did not reflect reliable penetration data. Thus, effective protease inhibitor(s) should be used to obtain the data corresponding to the in vivo transdermal experiment. This methodology will provide a means to eliminate the confounding effect of metabolism in permeation experiments.

Aminopeptidases↗
PubMed1996

Amino acid sequence of peptidyl-prolyl isomerase a of Fusarium sporotrichioides.

Peptidyl-prolyl-cis-trans isomerase catalyzes the interconversion of the cis and trans isomers of the proline-containing polypeptides and the folding process of proteins. This protein was known to be cyclophilin which has high binding affinity for cyclosporin A, a cyclic undecapeptide of fungal origin with potent immunosuppressive property agent. The two cytosolic peptidyl-prolyl-isomerases were found from Fusarium sporotrichioides. The amino acid sequence of the major peptidyl-prolyl isomerase a was determined by conventional sequencing methods; the protein with a calculated molecular mass of 19.7 kDa consisting of 179 amino acids. The comparison of the amino acid sequence of peptidyl-prolyl-isomerase from Fusarium with that of Nucerospora crassa revealed a significant degree of amino acid sequence homology (82.2%).

Amino Acid Isomerases↗
PubMed1996

Two-dimensional electrophoresis of plant proteins and standardization of gel patterns.

Proteins of two plants, Arabidopsis thaliana and rice (Oryza sativa) were subjected to two-dimensional electrophoresis analysis with two modifications: (i) comigration of external standard marker proteins with resultant horizontal and vertical stripes in the gel, and (ii) deblocking with a vapor of aqueous heptafluorobutyric acid for N-acetylserine. Approximately 5000 protein spots were separated from both the five tissues of Arabidopsis and the nine tissues of rice. Over one hundred spots were electroblotted for N-terminal sequencing. Among the newly sequenced proteins, 62 were from Arabidopsis and 51 from rice.

Amino Acid Sequence↗
PubMed1995

Isolation and characterization of a Photosystem II complex from the red alga Cyanidium caldarium: association of cytochrome c-550 and a 12 kDa protein with the complex.

A Photosystem II (PS II) complex was purified from an acidophilic as well as a thermophilic red alga, Cyanidium caldarium. The purified PS II complex was essentially devoid of phycobiliproteins and other contaminating components, and showed a high oxygen-evolving activity of 2375 mumol O2/mg Chl per h using phenyl-p-benzoquinone as the electron acceptor. The expression of this high activity did not require addition of exogenous Ca2+, although EDTA reduced the activity by 40%. This effect of EDTA can be reversed not only by Ca2+ but also by Mg2+; a similar Mg2+ effect has been observed in purified cyanobacterial PS II but not in higher plant PS II. Immunoblotting analysis indicated the presence of major intrinsic polypeptides commonly found in PS II from cyanobacteria and higher plants as well as the extrinsic 33 kDa protein. Antibodies against the extrinsic 23 and 17 kDa proteins of higher plant PS II, however, did not crossreact with any polypeptides in the purified PS II, indicating the absence of these proteins in the red alga. In contrast, two other extrinsic proteins of 17 and 12 kDa were present in the red algal PS II; they were released by 1 M Tris or Urea/NaCl treatment but not by 1 M NaCl. The 17 kDa polypeptide was identified to be cytochrome c-550 from heme-staining, immunoblot analysis and N-terminal amino acid sequencing, and the 12 kDa protein was found to be homologous to the 12 kDa extrinsic protein of cyanobacterial PS II from its N-terminal sequence. These results indicate that PS II from the red alga is closely related to PS II from cyanobacteria rather than to that from higher plants, and that the replacement of PS II extrinsic cytochrome c-550 and the 12 kDa protein by the extrinsic 23 and 17 kDa proteins occurred during evolution from red algae to green algae and higher plants.

Amino Acid Sequence↗
PubMed1995

Amino acid sequence of spinach ferredoxin:thioredoxin reductase catalytic subunit and identification of thiol groups constituting a redox-active disulfide and a [4Fe-4S] cluster.

Ferredoxin:thioredoxin reductase is a [4Fe-4S] protein involved in the light regulation of carbon metabolism in oxygenic photosynthesis. This enzyme catalyses the reduction of thioredoxins with light-generated electrons. Ferredoxin:thioredoxin reductase is composed of two dissimilar subunits, a catalytic subunit, and a variable subunit. The catalytic subunit of spinach ferredoxin:thioredoxin reductase, which contains the redox-active disulfide bridge, was sequenced by conventional protein sequencing techniques and the functional roles of all eight cysteine residues were examined by chemical modifications. The polypeptide chain with a calculated molecular mass of 12,959 Da consists of 113 amino acids and has a calculated isoelectric point of 5.30. Six of the eight cysteine residues are clustered as Cys-Pro-Cys and Cys-His-Cys groups. Cys19 and Cys27 are free cysteines with no catalytic function, Cys54 and Cys84 constitute the redox-active disulfide bridge of the active site, and the remaining four, Cys52, Cys71, Cys73, and Cys82 bind the Fe-S cluster.

Amino Acid Sequence↗
PubMed1995

Carboxy-terminal degradation of peptides using perfluoroacyl anhydrides. A C-terminal sequencing method.

An accurate carboxy-terminal sequencing method has long been sought to complement the Edman degradation procedure for amino-terminal amino acid sequence analysis. The method presented here is a unique and simple method to partly fulfill the needs. Exposure of a polypeptide to perfluoroacyl anhydride vapor at -20 degrees C for 0.5-1 h causes sequential chemical degradation of the molecule from the C-terminus. Fast-atom-bombardment mass spectrometric analysis of the resultant mixture of C-terminally truncated molecules permits the determination of the C-terminal sequence by simple calculation of the mass differences in molecular ions. Experiments suggested that this C-terminal degradation proceeds by active intermediates such as oxazolone at the C-terminal carboxyl residues.

Acetic Anhydrides↗
PubMed1995

Separation and characterization of Arabidopsis thaliana proteins by two-dimensional gel electrophoresis.

Arabidopsis (Arabidopsis thaliana) proteins were isolated from five tissues (leaf, stem, root, seed and callus), and separated by two-dimensional gel electrophoresis (2-DE). 2-DE was carried out by immobilized pH gradient (IPG) in the first dimension, and by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) in the second dimension. With the aid of comigrated five-marker proteins, the patterns of 2-DE gels for each tissue were graphically combined by a computer into a single synthetic image for the integrated Arabidopsis protein spots. The protein spot images, altogether 4763, were characterized by both molecular mass and isoelectric point. Partial amino(N)-terminal sequences of 101 protein spots were analyzed by Edman degradation. Fifty seven proteins were partially sequenced and 46 proteins appeared to have blocked N-termini. Deblocking by hydrazine vapor was carried out on 14 proteins and two of them were found to be pyroglutamyl-blocked N-termini. Forty seven new proteins were found by the present investigation.

Amino Acid Sequence↗
PubMed1994

Amino acid sequence of spinach ferredoxin:thioredoxin reductase variable subunit.

Ferredoxin:thioredoxin reductase (FTR) is an iron-sulfur protein, which, in the presence of ferredoxin and thioredoxin, catalyses the light-dependent activation of several photosynthetic enzymes. Spinach FTR consists of two dissimilar polypeptide chains, A and B, present in equal amounts. Whereas subunit B seems to be responsible for the catalytic activity, subunit A has no known catalytic function. We found earlier that the N-terminus of subunit A, also called the variable subunit, shows terminal redundancy and that 2-3 of its serine residues are phosphorylated [Tsugita, A. Yano, K., Gardet-Salvi, L. & Schürmann, P. (1991) Protein Sequence Data Anal. 4, 9-13]. We now report the complete amino acid sequence of subunit A, determined by conventional protein sequencing methods. The polypeptide chain with a calculated molecular mass of 12,669 Da consists of 112 amino acids and has a calculated isoelectric point of 5.4. The analysis of the sequence supports the idea that this subunit has no catalytic function. The comparison with a known cyanobacterial FTR reveals about 58% similarity and the striking presence of a N-terminal extension in the spinach protein. This extension may be responsible for the reported size variability of this subunit.

Amino Acid Sequence↗
PubMed1994

Separation and characterization of rice proteins.

Rice proteins from nine tissues and one organelle (leaf, chloroplast, stem, root, germ, dark germinated seedling, seed, bran, chaff and callus) were isolated and then separated by two-dimensional gel electrophoresis (2-DE). The protein spots were characterized according to molecular weight, isoelectric point and partial amino-terminal sequence. Electrophoresis was carried out by isoelectric focusing (IEF), nonequilibrium pH gradient electrophoresis (NEPHGE) and immobilized pH gradient (IPG) in the first dimension, and by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) in the second dimension. With the aid of nine marker proteins, the patterns of IEF, NEPHGE and IPG 2-DE gels were graphically combined by computer into a single synthetic image for each tissue, respectively, and these images for the nine tissues and one organelle were again combined into a single 2-DE image for the integrated rice protein spots. The rice 2-DE gel image resolved 4892 proteins. About 3% of the spots are characterized by amino-terminal sequencing.

Amino Acid Sequence↗
PubMed1994

Specific chemical cleavage of asparaginyl and glycyl-glycine bonds in peptides and proteins by anhydrous hydrazine vapor.

Hydrazinolysis of peptide or protein has been used for C-terminal amino acid determination by Akabori et al. (1952). In this study, proteins were reacted with anhydrous hydrazine vapor at 20 degrees C for 16 h. Asparaginyl linkages were cleaved. Asparagine and glutamine were converted to their hydrazides, beta-hydrazidyl aspartic acid and gamma-hydrazidyl glutamic acid, respectively, even under milder conditions. The former hydrazide cyclizes to a 6-membered ring, asparaginyl bond at the carboxyl side. Other cleavages, including the glycyl-glycine bond, were also observed.

Amino Acid Sequence↗
PubMed1993

Removal of N-terminal formyl groups and deblocking of pyrrolidone carboxylic acid of proteins with anhydrous hydrazine vapor.

Many proteins have a blocked alpha-amino group which renders them inaccessible to sequence analysis by the classical Edman degradation procedure. Blockage typically occurs when the alpha-amino groups are acylated with acetyl or formyl groups or when the N-terminal residue is pyrrolidone carboxylic acid formed by cyclization of glutamine. We have found that N-formyl groups of proteins and peptides can be removed by exposure to hydrazine vapor at -5 degrees C for 8 h. Under these conditions, peptide-bond cleavage or modification of the constituent amino-acid residues does not occur. Deblocking of N-terminal pyrrolidone carboxylate residues by conversion to gamma-hydrazidyl glutamic acid can be achieved by exposure to hydrazine vapor at 20 degrees C for 4 h. These conditions cause partial modification of asparagine and glutamine residues to their corresponding hydrazides, and conversion of arginine residues to ornithine.

Amino Acid Sequence↗
PubMed1993

[An experimental and clinical study of chorioretinal photocoagulation using a frequency-doubled Nd: YAG laser].

We developed a new frequency-doubled Nd: YAG laser system which works on 100 volts AC and emits continuous wave green laser light of 532 nm. Histological observation of rabbit eyes revealed no difference in chorioretinal lesions produced with the frequency-doubled Nd: YAG laser or with an argon green laser. This device was then clinically applied to 69 eyes of 59 patients with several disorders such as diabetic retinopathy, retinal vein occlusion, Eales' disease, Coats' disease, age-related macular degeneration, lattice degeneration in the peripheral retina, macular hole, and retinal tear. The ophthalmoscopic findings showed no visible differences in the lesions made with this device or with the argon laser. No complications were observed in these patients except for one case of central retinal vein occlusion, which developed vitreous hemorrhage after the treatment. This solid-state laser system has some mechanical advantages over the gas laser, i.e. reduced cooling requirements, easy maintenance, and long life, and it is considered to be usable clinically.

Adolescent↗
PubMed1992

C-terminal sequencing of protein. A novel partial acid hydrolysis and analysis by mass spectrometry.

Peptides or proteins were hydrolyzed by vapors of 90% pentafluoropropionic acid or heptafluorobutyric acid at 90 degrees C for various time periods. The hydrolyzate mixtures analyzed by both fast-atom-bombardment and electrospray ionization mass spectrometry showed a series of C-terminal successive degradation molecular ions. The degradation reaction may be due to the selective formation of an oxazolone ring at the C-terminal amino acid, followed by hydrolytic removal of the C-terminal amino acid. The major side reactions were cleavages of the peptide bonds at the C side of the internal aspartic acid residue and the N side of serine residue.

Amino Acid Sequence↗
PubMed1992

Purification and characterization of the major cationic kallikrein inhibitor in bovine pituitary gland.

The presence of two types of kallikrein inhibitor (cationic and anionic inhibitors) was demonstrated in bovine pituitary gland. These kallikrein inhibitors were separated from the homogenate of bovine posterior pituitary by successive CM-Sephadex chromatography. The major cationic inhibitor was further purified to homogeneity by affinity chromatography using porcine pancreatic beta-kallikrein immobilized on Sepharose 4B and gel filtration. The complete amino acid sequence of this inhibitor was first determined, and it was shown to be a peptide of 58 residues with a calculated molecular weight of 6,511. The Ki value against bovine pituitary kallikrein was 6 x 10(-9) M. The cationic inhibitor was found to be identical with basic pancreatic trypsin inhibitor.

Amino Acid Sequence↗