Characterization of cytoplasmic fibril structures found in gliding cells of Saprospira sp.

2005 ◽  
Vol 51 (10) ◽  
pp. 875-880 ◽  
Author(s):  
Gou Furusawa ◽  
Takeshi Yoshikawa ◽  
Yoshitaka Takano ◽  
Kazuyuki Mise ◽  
Iwao Furusawa ◽  
...  
Keyword(s):  
Amino Acid ◽  
Protein Band ◽  
Amino Acid Sequences ◽  
Gliding Motility ◽  
Pcr Product ◽  
Nutrient Agar Medium ◽  
Sds Page ◽  
Tail Sheath ◽  
A Subunit ◽  
Triton X

The cytoplasmic fibril structures of Saprospira sp. strain SS98-5 grown on a low-nutrient agar medium were purified from cell lysates treated with Triton X-100 and were observed by electron microscopy to be about 7 nm in width and 200–300 nm in length. SDS–PAGE of the fibril structures exhibited a single protein band with a molecular mass of 61 kDa. A Saprospira cytoplasmic fibril protein (SCFP), which is a subunit of the fibril structures, was digested with trypsin to oligopeptides and analyzed for amino acid sequences. A partial nucleotide sequence of the SCFP gene was determined after PCR using primers designated from the amino acid sequences of the oligopeptides. SCFP gene including DNA fragments were detected by Southern hybridization using the PCR product for an SCFP gene as a probe and were cloned to determine whole nucleotide sequences. The SCFP gene indicated relatively higher similarity to conserved hypothetical phage tail sheath proteins. A Western immunoblotting analysis showed that SCFP was significantly expressed in gliding cells as compared with nongliding cells. The above findings with the previously reported results suggest that the cytoplasmic fibril structures are possibly related to the gliding motility of Saprospira sp. strain SS98-5.Key words: Saprospira, gliding motility, Saprospira cytoplasmic fibril protein (SCFP).

ISOLATION AND CHARACTERIZATION OF THE GENES FOR THE a AND b SUBUNITS OF HUMAN COAGULATION FACTOR XIII

1987 ◽  
Author(s):  
A Ichinose ◽  
R E Bottenus ◽  
K R Loeb ◽  
E W Davie
Keyword(s):  
Amino Acid ◽  
Factor Xiii ◽  
Coagulation Factor ◽  
Amino Acid Sequences ◽  
Binding Region ◽  
Recombinant Phage ◽  
B Subunit ◽  
Isolation And Characterization ◽  
A Subunit ◽  
B Subunits

Factor XIII (plasma transglutaminase, fibrin stabilizing factor) is a plasma protein that plays an important role in the final stages of blood coagulation and fibrinolysis. The molecule occurs in blood as a tetramer (a2b2) consisting of two a. subunits and two b subunits. Recently, we have determined the amino acid sequences for both the a. and b subunits of human factor XIII by a combination of cDNA cloning and amino acid sequence analysis. cDNAs coding for the a (3.8 Kb) and b (2.2 Kb) subunits were used for the screening of human genomic DNA libraries. Among 12 × 106 recombinant phage, ∼30 have been shown to contain the sequences for the a subunit and ∼10 have been shown to contain the gene for the b subunit of factor XIII. The clones coding for the a. subunit span ∼90 Kb and have been characterized by restriction mapping. Southern blotting, and DNA sequencing. Both 5’ and 3’ ends of the genomic clones correspond to the 5’ and 3’portions of the cDNA for the a.subunit of factor XIII. The DNA sequence revealed that the activation peptide released ^thrombin (amino acid residues 137), the first putative Ca2+ binding region (around residue 251), the active Site Cys (amino acid residue 314), and the second putative Ca2+ binding region (around residue 473) are encoded by separate exons. Accordingly, the intervening sequences may separate the a subunit into functional and structural domains. The gene organization for the b subunit will also be presented. (Supported by NIH Grant HL 16919.)

Biochemical and genetic characterisation of a D glutenin subunit encoded at the Glu-B3 locus

Genome ◽  
1998 ◽  
Vol 41 (2) ◽  
pp. 215-220 ◽  
Author(s):  
M T Nieto-Taladriz ◽  
M Rodríguez-Quijano ◽  
J M Carrillo
Keyword(s):  
Amino Acid ◽  
Amino Acid Sequence ◽  
Wheat Gluten ◽  
Gene Clusters ◽  
Glutenin Subunits ◽  
Key Words Wheat ◽  
Sds Page ◽  
A Subunit ◽  
Omega Gliadins ◽  
Prolamin Loci

The SDS-PAGE pattern of reduced and alkylated glutenins from the bread wheat cultivar Prinqual presents a subunit (named d4) in the mobility zone of the omega -gliadins that only appears under reduced conditions. This subunit was isolated and characterised at the biochemical and genetic levels. Subunit d4 was shown to form disulphide aggregates with glutenins and had an acidic pI. These characteristics correspond to those of the D glutenin subunits. The N-terminal amino acid sequence of subunit d4 was coincident with the SRL sequence type characteristic of omega -gliadins encoded by genes on the 1B chromosome, and confirms the similarity between D glutenin subunits and omega -gliadins. The genetic study of subunit d4 was performed in the F2 progeny from the 'Prinqual' x 'Ablaca' cross, based on four prolamin loci: Glu-B1, Glu-B3, Gli-B1, and Gli-B5. The recombinants found between Glu-B3 and Gli-B1 demonstrated that subunit d4 was encoded at the Glu-B3 locus, and reinforces the hypothesis of the duplication of prolamin gene clusters in wheat. A preliminary study of the effect of subunit d4 on gluten strength showed that lines with the Glu-B3 allele from 'Prinqual', which includes subunit d4, had a significantly higher sedimentation volume than those with the allele from 'Ablaca'.Key words: wheat gluten proteins, D glutenin subunits, amino acid sequence, linkage mapping, complex loci duplication.

scholarly journals Sulphoacetaldehyde sulpho-lyase (EC 4.4.1.12) from Desulfonispora thiosulfatigenes: purification, properties and primary sequence

2001 ◽  
Vol 357 (2) ◽  
pp. 581-586 ◽  
Author(s):  
Karin DENGER ◽  
Jürgen RUFF ◽  
Ulrike REIN ◽  
Alasdair M. COOK
Keyword(s):  
Amino Acid ◽  
Molecular Mass ◽  
Pcr Primers ◽  
Amino Acid Sequences ◽  
Thiamine Pyrophosphate ◽  
Strictly Anaerobic ◽  
Laser Desorption Ionization ◽  
Native Form ◽  
Ionization Time ◽  
Sds Page

The strictly anaerobic bacterium Desulfonispora thiosulfatigenes ferments taurine via sulphoacetaldehyde, which is hydrolysed to acetate and sulphite by sulphoacetaldehyde sulpho-lyase (EC 4.4.1.12). The lyase was expressed at high levels and a two-step, 4.5-fold purification yielded an apparently homogeneous soluble protein, which was presumably a homodimer in its native form; the molecular mass of the subunit was about 61kDa (by SDS/PAGE). The mass was determined to be 63.8kDa by matrix-assisted laser-desorption ionization–time-of-flight (MALDI–TOF) MS. The purified enzyme converted 1mol of sulphoacetaldehyde to 1mol each of sulphite and acetate, but no requirement for thiamine pyrophosphate (TPP) was detected. The N-terminal and two internal amino acid sequences were determined, which allowed us to generate PCR primers. The gene was amplified and sequenced. The DNA sequence had no significant homologue in the databases searched, whereas the derived amino acid sequence indicated an oxo-acid lyase, revealed a TPP-binding site and gave a derived molecular mass of 63.8kDa.

scholarly journals Equimolar heterodimers in microtubules.

1982 ◽  
Vol 94 (2) ◽  
pp. 263-270 ◽  
Author(s):  
R E Stephens
Keyword(s):  
Amino Acid ◽  
Tryptic Peptide ◽  
Optimal Conditions ◽  
Beta Chain ◽  
Closely Related Species ◽  
Sds Page ◽  
Amino Acid Profiles ◽  
Triton X ◽  
Brain Tubulin ◽  
Sea Urchin Sperm

Two equimolar beta chains can be resolved from sea urchin sperm flagellar and scallop gill ciliary tubulins, and from certain brain tubulins as well, using the Triton X-100-acid-urea polyacrylamide gel system commonly used for histone analysis. The beta chains are identified as such from their mobility on urea-free SDS PAGE, from amino acid composition, and from tryptic peptide distribution. Scallop beta chains have almost identical amino acid profiles but they differ by one tryptic peptide. Optimal conditions for beta chain resolution are very species-dependent, with some closely related species showing either maximal or no beta chain separation. In addition, beef brain tubulin on Triton X-100-acid-urea electrophoresis and scallop gill ciliary tubulin upon isoelectric focusing in the presence of SDS show two approximately equimolar alpha chains. These data, indicating equimolar amounts of two potentially different tubulin heterodimers from a variety of microtubule types, support a model for microtubule structure wherein protofilaments consist of alternating heterodimers of two kinds, generating a 16-nm (2-dimer) axial repeat.

scholarly journals Conformational differences between two wheat (Triticum aestivum) ‘high-molecular-weight’ glutenin subunits are due to a short region containing six amino acid differences

1989 ◽  
Vol 263 (3) ◽  
pp. 837-842 ◽  
Author(s):  
A P Goldsbrough ◽  
N J Bulleid ◽  
R B Freedman ◽  
R B Flavell
Keyword(s):  
Molecular Weight ◽  
Triticum Aestivum ◽  
Amino Acid ◽  
High Molecular Weight ◽  
Polyacrylamide Gel Electrophoresis ◽  
Amino Acid Sequences ◽  
Glutenin Subunits ◽  
Sds Page ◽  
Hmw Glutenin

‘High-molecular-weight’ (HMW, high-Mr) glutenin subunits are protein constituents of wheat (Triticum aestivum) seeds and are responsible in part for the viscoelasticity of the dough used to make bread. Two subunits, numbered 10 and 12, are the products of allelic genes. Their amino acid sequences have been derived from the nucleic acid sequences of the respective genes. Subunit 10 has fewer amino acids than subunit 12, but migrates more slowly on SDS/PAGE (polyacrylamide-gel electrophoresis). This anomaly is due to between one and six of the amino acid differences between the subunits, localized towards the C-terminal end of the proteins. This has been established by making chimaeric genes between the genes for subunits 10 and 12, transcribing and translating them in vitro and analysing the products by SDS/PAGE. The postulated conformational differences between subunits 10 and 12 are discussed in relation to current hypotheses for the structure of HMW glutenin subunits.

scholarly journals Affinity purification of 5-methylthioribose kinase and 5-methylthioadenosine/S-adenosylhomocysteine nucleosidase from Klebsiella pneumoniae

1996 ◽  
Vol 317 (1) ◽  
pp. 285-290 ◽  
Author(s):  
Kenneth A. CORNELL ◽  
R. W. WINTER ◽  
Paula A. TOWER ◽  
Michael K. RISCOE
Keyword(s):  
Amino Acid ◽  
Molecular Mass ◽  
Sequence Similarity ◽  
Affinity Purification ◽  
Amino Acid Sequences ◽  
Reading Frame ◽  
Sds Page ◽  
High Degree ◽  
Putative Translation Product ◽  
Substrate Kinetics

Two enzymes in the methionine salvage pathway, 5-methylthioribose kinase (MTR kinase) and 5´-methylthioadenosine/S-adenosylhomocysteine nucleosidase (MTA/SAH nucleosidase) were purified from Klebsiellapneumoniae. Chromatography using a novel 5´-(p-aminophenyl)thioadenosine/5-(p-aminophenyl)thioribose affinity matrix allowed the binding and selective elution of each of the enzymes in pure form. The molecular mass, substrate kinetics and N-terminal amino acid sequences were characterized for each of the enzymes. Purified MTR kinase exhibits an apparent molecular mass of 46–50 kDa by SDS/PAGE and S200HR chromatography, and has a Km for MTR of 12.2 μM. Homogeneous MTA/SAH nucleosidase displays a molecular mass of 26.5 kDa by SDS/PAGE, and a Km for MTA of 8.7 μM. Comparisons of the N-terminal sequences obtained for each of the enzymes with protein-sequence databases failed to reveal any significant sequence similarities to known proteins. However, the amino acid sequence obtained for the nucleosidase did share a high degree of sequence similarity with the putative translation product of an open reading frame in Escherichia coli, thus providing a tentative identification of this gene as encoding an MTA/SAH nucleosidase.

scholarly journals THE CIDR1α-PfEMP1 SEQUENCE FROM INDONESIAN PLASMODIUM FALCIPARUM AND ITS POTENTIAL ASSOCIATION WITH THE CEREBRAL OUTCOME

2021 ◽  
Vol 7 (1) ◽  
pp. 34-39
Author(s):  
Erma Sulistyaningsih ◽  
Yunita Armiyanti ◽  
Rosita Dewi
Keyword(s):  
Plasmodium Falciparum ◽  
Amino Acid ◽  
Cerebral Malaria ◽  
Severe Malaria ◽  
Sequence Similarity ◽  
Malaria Diagnosis ◽  
Severe Case ◽  
Amino Acid Sequences ◽  
Pcr Product ◽  
Potential Association

Background: Plasmodium falciparum Erythrocyte Membrane Protein 1 (PfEMP1) is an important protein responsible for the pathogenesis of severe malaria, including cerebral malaria. The protein is highly diverse. The CIDR1α-PfEMP1 binds endothelial protein receptor (EPCR) and may associated with the brain swelling in childhood malaria. Objective: To analyze the CIDR1α-PfEMP1 from Indonesian isolate and determine its association with cerebral malaria outcome. Methods: Fifteen blood samples of clinically mild to severe malaria-patient were collected for DNA extraction. Malaria diagnosis was conducted microscopically by Giemsa-stained thin blood smear. The CIDR1α domain was amplified by PCR using specific primer and PCR product was sequenced. The nucleotide sequences were analyzed by NCBI blast, DNASIS MAX 3 and translated into amino acid sequences using Expasy Translation Tool. Results: One out of fifteen samples was severe malaria case and infected with P. falciparum, the rest were clinically mild to moderate malaria and infected with pure P. falciparum or mixed infection of P. falciparum and P. vivax. Amplification for CIDR1α domain resulted a single band of + 550 bp from a severe sample only. Sequencing of PCR product on both strands read 524 nucleotides and BLAST analysis confirmed as CIDR1α sequence. Multiple alignment showed 74-78% nucleotide sequence similarity with reference sequences, but amino acid sequences presented 23.5% homologous. Conclusion: An identified CIDR1α domain only from severe case implicating the potential association with the severe outcome including cerebral malaria, but the highly diverse of the domain needs further studies on the interaction with the pathological-causing receptor in the host.

scholarly journals β-Galactosidase II in Ripening Tomatoes

HortScience ◽  
1995 ◽  
Vol 30 (4) ◽  
pp. 817A-817
Author(s):  
Russell Pressey ◽  
C.M. Sean Carrington
Keyword(s):  
Amino Acid ◽  
Cell Walls ◽  
Amino Acid Sequences ◽  
Cell Wall Polysaccharides ◽  
Terminal Amino Acid ◽  
Molecular Weights ◽  
Sds Page ◽  
Original Procedure ◽  
Terminal Amino ◽  
Terminal Amino Acid Sequence

Tomatoes contain several isozymes of β-galactosidase, but only one, β-galactosidase II, can hydrolyze the β-1,4-galactans in tomato cell walls. β-galactosidase II has now been highly purified by modification of the original procedure. The molecular weight of this isozyme is ≈62 kDa according to gel infiltration, but SDS-PAGE of the purified enzyme separated three components with molecular weights of 29, 42, and 82 kDa. The 82-kDa peptide may be the intact enzyme and the smallest peptides are subunits as proposed for other β-galactosidases. The N-terminal amino acid sequence of β-galactosidase II showed high homology with amino acid sequences reported for other plant β-galactosidases. A new assay for β-galactosidase II in tomato extracts has been developed using FPLC. This isozyme was not detected in mature-green tomatoes but appeared at about the breaker stage and increased during ripening. The increase in b-galactosidase II was accompanied by a decrease in galactose content of cell wall polysaccharides, suggesting that this enzyme may be involved in the loss of galactose during tomato ripening.

Identification of thecrygene inBacillus thuringiensisstrain WZ-9 and its toxicity againstHenosepilachna vigintioctomaculata

2008 ◽  
Vol 5 (3) ◽  
pp. 245-250 ◽  
Author(s):  
Song Ping ◽  
Wang Qin-Ying ◽  
Wu Hui-Xian ◽  
Lu Xiu-Jun ◽  
Wang Yong
Keyword(s):  
Amino Acid ◽  
Culture Media ◽  
Stationary Phases ◽  
Protein Band ◽  
Growth Cycle ◽  
Amino Acid Residues ◽  
Reading Frame ◽  
Gene Sequence Analysis ◽  
Sds Page ◽  
Encoding Gene

AbstractBacillus thuringiensisstrain WZ-9, isolated from soil in Hebei province, China, was effective againstHenosepilachna vigintioctomaculatalarvae. The strain presented bipyramidal crystals with a protein band of 130 kDa in SDS–PAGE. The pH changes of the culture media showed important fluctuations during the 24 h growth cycle. The pH varied less in log and stationary phases than it did in the exponential phase. Bioassay results showed that the WZ-9 strain was only harmful to larvae ofH. vigintioctomaculataand not to either adults ofH. vigintioctomaculataor other several lepidopteran and coleopteran insects. LC50to second-instar larvae ofH. vigintioctomaculatawas 2.95×107cells/ml after 72 h. Genotypic investigations showed that this strain possessed thecry7gene. Sequence analysis demonstrated that the encoding gene contained an open reading frame (ORF) of 3414 bp and encoded 1138 amino acid residues. The deduced amino acid sequence was 99.65% identical to that of the reported Cry7Ab2 sequences. This gene was designated by the Bt δ-endotoxin nomenclature committee as Cry7Ab3 with accession number BI 1015188 in the GenBank database.

scholarly journals The primary structure of bovine monoamine oxidase type A. Comparison with peptide sequences of bovine monoamine oxidase type B and other flavoenzymes

1989 ◽  
Vol 259 (2) ◽  
pp. 407-413 ◽  
Author(s):  
J F Powell ◽  
Y P Hsu ◽  
W Weyler ◽  
S Chen ◽  
J Salach ◽  
...  
Keyword(s):  
Amino Acid ◽  
Monoamine Oxidase ◽  
Type A ◽  
Amino Acid Sequences ◽  
Cdna Clones ◽  
Amino Acid Residues ◽  
Type B ◽  
Sequence Comparisons ◽  
A Subunit ◽  
Flavin Binding

We have isolated cDNA clones believed to encompass the full-length coding sequences for a subunit of bovine monoamine oxidase type A (MAO-A). The clones code for an apoprotein of 527 amino acid residues corresponding to a molecular mass of 59,806 Da. The inferred protein sequences show an overall similarity of 68% with partial amino acid sequences of bovine type B MAO (about 41% of the total sequence), as well as a greater similarity (greater than 90%) with some regions including that for the published sequence of the flavin-binding region. Sequence comparisons indicate that these two forms of MAO are encoded by distinct genes. Comparison of this sequence with other flavoenzymes showed similarity with regions associated with non-covalent flavin-binding sites. Analysis of mRNAs coding for MAO enzymes showed a heterogeneity of transcripts consistent with several different forms of monoamine oxidase.

Sign in / Sign up

Export Citation Format

Share Document