scholarly journals The Presence and Characterization of a virF Gene on Agrobacterium vitis Ti Plasmids

1998 ◽  
Vol 11 (5) ◽  
pp. 429-433 ◽  
Author(s):  
B. Schrammeijer ◽  
J. Hemelaar ◽  
P. J. J. Hooykaas
Keyword(s):  
Amino Acids ◽  
Agrobacterium Tumefaciens ◽  
Molecular Mass ◽  
Promoter Region ◽  
Consensus Sequence ◽  
Tumor Formation ◽  
Nicotiana Glauca ◽  
Agrobacterium Vitis ◽  
Ti Plasmids

Octopine and nopaline strains of Agrobacterium tumefaciens differ in their ability to induce tumors on Nicotiana glauca. The presence of a virF locus on the octopine Ti plasmid makes N. glauca a host plant for these strains, indicating that the VirF protein is a host-range determinant. Here we show the presence of a virF locus not only on the Agrobacterium vitis octopine/cucumopine plasmids pTiAg57 and pTiTm4, but also on the nopaline Ti plas-mids pTiAT1, pTiAT66a, and pTiAT66b. On the octopine Ti plasmids from A. tumefaciens the virF gene is located between the virE locus and the left border of the T-region. In contrast, the virF gene on Ti plasmids of A. vitis is located at the very left end of the vir-region near the virA locus. The virF gene of pTiAg57 has been sequenced and codes for a protein of 202 amino acids with a molecular mass of 22,280 Da. Comparison showed that the virF gene from A. vitis strain Ag57 is almost identical to that from A. tumefaciens octopine strains. The transcription of the pTiAg57 virF is inducible by the plant phenolic compound acetosyringone through the presence of a vir-box consensus sequence in its promoter region. The VirF protein from pTiAg57 can complement octopine A. tumefaciens strains deleted for virF as shown by tumor formation on N. glauca.

scholarly journals Characterization of an Extracellular Dipeptidase from Streptococcus gordonii FSS2

2005 ◽  
Vol 73 (2) ◽  
pp. 1256-1259 ◽  
Author(s):  
J. M. Goldstein ◽  
T. Kordula ◽  
J. L. Moon ◽  
J. A. Mayo ◽  
J. Travis
Keyword(s):  
Amino Acids ◽  
Gene Family ◽  
Molecular Mass ◽  
Streptococcus Gordonii ◽  
Its Gene ◽  
Theoretical Molecular Mass

ABSTRACT PepV, a dipeptidase found in culture fluids of Streptococcus gordonii FSS2, was purified and characterized, and its gene was cloned. PepV is a monomeric metalloenzyme of approximately 55 kDa that preferentially degrades hydrophobic dipeptides. The gene encodes a polypeptide of 467 amino acids, with a theoretical molecular mass of 51,114 Da and a calculated pI of 4.8. The S. gordonii PepV gene is homologous to the PepV gene family from Lactobacillus and Lactococcus spp.

scholarly journals Mass Spectrometry Analysis and Biological Characterization of the Predatory Ant Odontomachus monticola Venom and Venom Sac Components

Toxins ◽  
2019 ◽  
Vol 11 (1) ◽  
pp. 50 ◽  
Author(s):  
Naoki Tani ◽  
Kohei Kazuma ◽  
Yukio Ohtsuka ◽  
Yasushi Shigeri ◽  
Keiichi Masuko ◽  
...  
Keyword(s):  
Mass Spectrometry ◽  
Amino Acids ◽  
Molecular Mass ◽  
Biogenic Amines ◽  
Transcriptome Analysis ◽  
Mass Spectrometry Analysis ◽  
Biological Characterization ◽  
Spectrometry Analysis ◽  
Dipeptidyl Peptidases

We previously identified 92 toxin-like peptides and proteins, including pilosulin-like peptides 1–6 from the predatory ant Odontomachus monticola, by transcriptome analysis. Here, to further characterize venom components, we analyzed the venom and venom sac extract by ESI-MS/MS with or without trypsin digestion and reducing agent. As the low-molecular-mass components, we found amino acids (leucine/isoleucine, phenylalanine, and tryptophan) and biogenic amines (histamine and tyramine) in the venom and venom sac extract. As the higher molecular mass components, we found peptides and proteins such as pilosulin-like peptides, phospholipase A2s, hyaluronidase, venom dipeptidyl peptidases, conotoxin-like peptide, and icarapin-like peptide. In addition to pilosulin-like peptides 1–6, we found three novel pilosulin-like peptides that were overlooked by transcriptome analysis. Moreover, pilosulin-like peptides 1–6 were chemically synthesized, and some of them displayed antimicrobial, hemolytic, and histamine-releasing activities.

scholarly journals Characterization of an Agrobacterium tumefaciensd-Psicose 3-Epimerase That Converts d-Fructose to d-Psicose

2006 ◽  
Vol 72 (2) ◽  
pp. 981-985 ◽  
Author(s):  
Hye-Jung Kim ◽  
Eun-Kyung Hyun ◽  
Yeong-Su Kim ◽  
Yong-Joo Lee ◽  
Deok-Kun Oh
Keyword(s):  
Escherichia Coli ◽  
Agrobacterium Tumefaciens ◽  
Molecular Mass ◽  
Specific Activity ◽  
Catalytic Efficiency ◽  
Maximal Activity ◽  
Turnover Number ◽  
Conversion Yield ◽  
Equilibrium Ratio

ABSTRACT The noncharacterized gene previously proposed as the d-tagatose 3-epimerase gene from Agrobacterium tumefaciens was cloned and expressed in Escherichia coli. The expressed enzyme was purified by three-step chromatography with a final specific activity of 8.89 U/mg. The molecular mass of the purified protein was estimated to be 132 kDa of four identical subunits. Mn2+ significantly increased the epimerization rate from d-fructose to d-psicose. The enzyme exhibited maximal activity at 50°C and pH 8.0 with Mn2+. The turnover number (k cat) and catalytic efficiency (k cat/Km ) of the enzyme for d-psicose were markedly higher than those for d-tagatose, suggesting that the enzyme is not d-tagatose 3-epimerase but d-psicose 3-epimerase. The equilibrium ratio between d-psicose and d-fructose was 32:68 at 30°C. d-Psicose was produced at 230 g/liter from 700-g/liter d-fructose at 50°C after 100 min, corresponding to a conversion yield of 32.9%.

scholarly journals Cloning and Characterization of a Tetracycline Resistance Determinant Present in Agrobacterium tumefaciens C58

1999 ◽  
Vol 181 (2) ◽  
pp. 618-626 ◽  
Author(s):  
Zhao-Qing Luo ◽  
Stephen K. Farrand
Keyword(s):  
Agrobacterium Tumefaciens ◽  
Promoter Region ◽  
Tetracycline Resistance ◽  
Homologous Region ◽  
Polycyclic Compounds ◽  
Repressor Proteins ◽  
Repressive Effect ◽  
Agrobacterium Tumefaciens C58 ◽  
Spontaneous Mutants

ABSTRACT Agrobacterium tumefaciens C58 and its derivatives give rise to spontaneous mutants resistant to tetracycline at a high frequency. We observed that a mutation affecting a tRNA processing function significantly affected the emergence of such mutants, suggesting that C58 contained a positively acting gene conferring resistance to tetracycline. A cosmid clone conferring resistance to tetracycline in Escherichia coli andAgrobacterium was isolated from a genomic bank of one such mutant. Subcloning, transposon mutagenesis, and DNA sequence analysis revealed that this DNA fragment contained two divergently transcribed genes, tetA and tetR, encoding products that were very similar to proteins of the Tet(A) class of tetracycline resistance systems. In the clone from this mutant, tetR was disrupted by an IS426. The homologous region from wild-type NT1 contained an intact tetR gene and did not confer resistance to tetracycline. Hybridization analysis showed that of 22 members of the genus Agrobacterium surveyed, only strains C58 and T37 contained the tet determinant. Moreover, only these two strains mutated to resistance to this antibiotic. Unlike other Tet(A) systems, neither tetracycline nor a series of its derivatives induced the expression of this tet gene unit. Other polycyclic compounds, including many of plant origin, also did not induce this tet gene system. The divergent promoter region of this tet system contained a single inverted repeat element identical to one such operator repeat in the promoter region of the tet determinant from the IncP1α R plasmid RP4. TetR repressor proteins from the Agrobacterium tetsystem and from RP4 interacted with the heterologous operators. While the repressive effect of the TetR protein from strain C58 (TetRC58) on the tetA gene from strain RP4 (tetA RP4) was not relieved by tetracycline, repression of tetA C58 by TetRRP4was lifted by this antibiotic.

Isolation and characterization of a heat-soluble protein from pea (Pisum sativum) embryos

1995 ◽  
Vol 5 (3) ◽  
pp. 137-144 ◽  
Author(s):  
Pauline S. Russouw ◽  
Jill Farrant ◽  
Wolf Brandt ◽  
Dennis Maeder ◽  
George G. Lindsey
Keyword(s):  
Amino Acids ◽  
Ionic Strength ◽  
Pisum Sativum ◽  
Consensus Sequence ◽  
High Ionic Strength ◽  
Supernatant Fraction ◽  
Chick Pea ◽  
Isolation And Characterization ◽  
Helical Content

AbstractAn LEA-like protein has been isolated and characterized from pea (Pisum sativum) embryos. It is the most prevalent protein in a homogenate of pea axes heated for 10 min at 80°C and then centifuged for 10 min at 17000g(80°C supernatant fraction). It has a molecular mass of 11 kDa and is very rich in hydrophilic amino acids, notably aspartate, glutamate and glycine. The protein is not recognized by an antibody to the group II dehydrin C-terminal consensus sequence. Antibodies to the isolated protein recognize a number of larger proteins in the 80°C supernatant fraction of pea and other legumes (chick pea, soybean and bean) as well as gramineous seeds (wheat, maize and barley). The protein undergoes a substantial increase in α-helical content at high ionic strength but does not dimerize.

Isolation and characterization of diverse nopaline type Ti plasmids of Agrobacterium tumefaciens from Japan

1989 ◽  
Vol 152 (2) ◽  
pp. 119-124 ◽  
Author(s):  
H. Wabiko ◽  
M. Kagaya ◽  
I. Kodama ◽  
K. Masuda ◽  
Y. Kodama ◽  
...  
Keyword(s):  
Agrobacterium Tumefaciens ◽  
Isolation And Characterization ◽  
Ti Plasmids

Molecular cloning and characterization of the yellowtail GH gene and its promoter: a consensus sequence for teleost and avian Pit-1/GHF-1 binding sites

1996 ◽  
Vol 16 (1) ◽  
pp. 63-72 ◽  
Author(s):  
T Ohkubo ◽  
M Araki ◽  
M Tanaka ◽  
S Sudo ◽  
K Nakashima
Keyword(s):  
Promoter Region ◽  
Consensus Sequence ◽  
Transcription Start ◽  
Dominant Form ◽  
Nuclear Extracts ◽  
Gh Gene ◽  
Seriola Quinqueradiata ◽  
Gel Shift Assay ◽  
Flanking Region

ABSTRACT The gene and 5′ flanking promoter region for yellowtail (Seriola quinqueradiata) GH (yGH) have been cloned, sequenced and characterized. The yGH gene spans approximately 4·6 kb and consists of six exons and five introns, as has been observed with rainbow trout, Atlantic salmon and tilapia GH genes. This result suggests that the structure of six exons and five introns is a dominant form in fish GH genes. A typical TATA box exists 26 bp upstream from the transcription start site, and Pit-1/GHF-1 (Pit-1) binding site-homologous regions were found in the promoter region of the yGH gene. In a gel shift assay, however, a single shifted band was detected with the fragments containing a region from −128 to −90 of the yGH 5′ flanking region when they were incubated with yellowtail pituitary nuclear extracts. The bound fragments contained an octamer base sequence similar, but not identical, to mammalian consensus Pit-1 binding element. A consensus octamer sequence is also proposed in this report for the binding of teleost and avian Pit-1 transcription factors.

scholarly journals Molecular characterization of the Agrobacterium tumefaciens DNA transfer protein VirB6

Microbiology ◽  
2005 ◽  
Vol 151 (11) ◽  
pp. 3483-3492 ◽  
Author(s):  
Paul K. Judd ◽  
David Mahli ◽  
Anath Das
Keyword(s):  
Amino Acids ◽  
Agrobacterium Tumefaciens ◽  
Transmembrane Domain ◽  
Ti Plasmid ◽  
Dna Transfer ◽  
Single Amino Acid ◽  
Dependent Manner ◽  
Cell Pole ◽  
A Cell

The VirB proteins of Agrobacterium tumefaciens assemble a T-pilus and a type IV secretion (T4S) apparatus for the transfer of DNA and proteins to plant cells. VirB6 is essential for DNA transfer and is a polytopic integral membrane protein with at least four membrane-spanning domains. VirB6 is postulated to function in T-pilus biogenesis and to be a component of the T4S apparatus. To identify amino acids required for VirB6 function, random mutations were introduced into virB6, and mutants that failed to complement a deletion in virB6 in tumour formation assays were isolated. Twenty-one non-functional mutants were identified, eleven of which had a point mutation that led to a substitution in a single amino acid. Characterization of the mutants indicated that the N-terminal large periplasmic domain and the transmembrane domain TM3 are required for VirB6 function. TM3 has an unusual sequence feature in that it is rich in bulky hydrophobic amino acids. This feature is found conserved in the VirB6 family of proteins. Studies on the effect of VirB6 on other VirB proteins showed that the octopine Ti-plasmid VirB6, unlike its nopaline Ti-plasmid counterpart, does not affect accumulation of VirB3 and VirB5, but has a strong negative effect on the accumulation of the VirB7-VirB7 dimer. Using indirect immunofluorescence microscopy the authors recently demonstrated that VirB6 localizes to a cell pole in a VirB-dependent manner. Mutations identified in the present study did not affect polar localization of the protein or the formation of the VirB7-VirB7 dimer. A VirB6-GFP fusion that contained the entire VirB6 ORF did not localize to a cell pole in either the presence or the absence of the other VirB proteins. IMF studies using dual labelling demonstrated that VirB6 colocalizes with VirB3 and VirB9, and not with VirB4, VirB5 and VirB11. These results support the conclusion that VirB6 is a structural component of the T4S apparatus.

scholarly journals Cloning and characterization of the cDNA coding for a polymyositis-scleroderma overlap syndrome-related nucleolar 100-kD protein.

1992 ◽  
Vol 176 (4) ◽  
pp. 973-980 ◽  
Author(s):  
M Blüthner ◽  
F A Bautz
Keyword(s):  
Amino Acids ◽  
Amino Acid ◽  
Protein Sequence ◽  
Epitope Mapping ◽  
Consensus Sequence ◽  
Overlap Syndrome ◽  
Amino Acid Residues ◽  
Protein Sequence Analysis ◽  
Protein Fragment

About 50% of patients with the polymyositis-scleroderma overlap syndrome are reported to have autoantibodies to a nucleolar particle termed PM/Scl. The particle consists of several polypeptides of which two proteins of 75 and 100 kD have been identified as the major antigenic components. Here we report on the cDNA cloning and partial epitope mapping of the 100-kD autoantigen from human placenta and HeLa lambda gt11 libraries. The deduced amino acid sequence encodes a protein of 885 amino acid residues with a molecular mass of 100.8 kD. Rabbit antibodies raised against a recombinant protein fragment reacted in immunofluorescence and immunoblotting in the same manner as human autoantibodies directed against the nucleolar 100-kD protein. Sequence analysis shows close homology to a consensus sequence of 12 amino acids from serine/threonine kinases, suggesting a possible function for this autoantigen. A major antigenic region is found to be located within the NH2-terminal third of the polypeptide.

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