scholarly journals Molecular cloning and characterization of ornithine decarboxylase cDNA of the nematode Panagrellus redivivus

1995 ◽  
Vol 308 (2) ◽  
pp. 635-640 ◽  
Author(s):  
H von Besser ◽  
G Niemann ◽  
B Domdey ◽  
R D Walter
Keyword(s):  
Amino Acids ◽  
Active Sites ◽  
Cdna Clones ◽  
Degenerate Primers ◽  
Calculated Molecular Mass ◽  
Panagrellus Redivivus ◽  
Free Living Nematode ◽  
Mixed Stage ◽  
Conserved Amino Acids

In a PCR with degenerate primers encoding highly conserved amino acids within ornithine decarboxylases (ODCs) of several organisms, a fragment of the ODC gene of the free-living nematode Panagrellus redivivus was isolated. Northern blot analysis revealed a single 1.7 kb transcript in a mixed-stage population of animals. From this RNA source, a cDNA library was constructed and screened with the PCR fragment. Several cDNA clones were isolated, one of which encodes the complete 435-amino-acid ODC enzyme with a calculated molecular mass of 47.1 kDa. The P. redivivus ODC possesses 126 of the 136 highly conserved amino acids in the enzymes from fungi, invertebrates and vertebrates. Functional amino acids are conserved, suggesting that the two active sites of the P. redivivus ODC are formed at the interface of a homodimer, as described for mammalian ODCs.

scholarly journals Molecular Characterization of a Novel Glucosyltransferase from Lactobacillus reuteri Strain 121 Synthesizing a Unique, Highly Branched Glucan with α-(1→4) and α-(1→6) Glucosidic Bonds

2002 ◽  
Vol 68 (9) ◽  
pp. 4283-4291 ◽  
Author(s):  
S. Kralj ◽  
G. H. van Geel-Schutten ◽  
H. Rahaoui ◽  
R. J. Leer ◽  
E. J. Faber ◽  
...  
Keyword(s):  
Amino Acids ◽  
Amino Acid ◽  
Amino Acid Sequence ◽  
Molecular Characterization ◽  
Lactobacillus Reuteri ◽  
Degenerate Primers ◽  
Terminal Domain ◽  
Branching Points ◽  
Culture Supernatants

ABSTRACT Lactobacillus reuteri strain 121 produces a unique, highly branched, soluble glucan in which the majority of the linkages are of the α-(1→4) glucosidic type. The glucan also contains α-(1→6)-linked glucosyl units and 4,6-disubstituted α-glucosyl units at the branching points. Using degenerate primers, based on the amino acid sequences of conserved regions from known glucosyltransferase (gtf) genes from lactic acid bacteria, the L. reuteri strain 121 glucosyltransferase gene (gtfA) was isolated. The gtfA open reading frame (ORF) was 5,343 bp, and it encodes a protein of 1,781 amino acids with a deduced M r of 198,637. The deduced amino acid sequence of GTFA revealed clear similarities with other glucosyltransferases. GTFA has a relatively large variable N-terminal domain (702 amino acids) with five unique repeats and a relatively short C-terminal domain (267 amino acids). The gtfA gene was expressed in Escherichia coli, yielding an active GTFA enzyme. With respect to binding type and size distribution, the recombinant GTFA enzyme and the L. reuteri strain 121 culture supernatants synthesized identical glucan polymers. Furthermore, the deduced amino acid sequence of the gtfA ORF and the N-terminal amino acid sequence of the glucosyltransferase isolated from culture supernatants of L. reuteri strain 121 were the same. GTFA is thus responsible for the synthesis of the unique glucan polymer in L. reuteri strain 121. This is the first report on the molecular characterization of a glucosyltransferase from a Lactobacillus strain.

The Drosophila gene asteroid encodes a novel protein and displays dosage-sensitive interactions with Star and Egfr

Genome ◽  
1998 ◽  
Vol 41 (2) ◽  
pp. 295-302 ◽  
Author(s):  
Michael A Kotarski ◽  
Deborah A Leonard ◽  
Sean A Bennett ◽  
Clifton P Bishop ◽  
Stephen D Wahn ◽  
...  
Keyword(s):  
Amino Acids ◽  
Egf Receptor ◽  
Open Reading Frame ◽  
Cdna Clones ◽  
Putative Open Reading Frame ◽  
Morphogenetic Furrow ◽  
Calculated Molecular Mass ◽  
Reading Frame ◽  
Ribonuclease Protection ◽  
Novel Protein

The asteroid gene of Drosophila was found to lie within 189 bp of Star. Asteroid cDNA clones were isolated and sequenced and a single putative open reading frame was identified that encodes a novel protein of 815 amino acids with a calculated molecular mass of 93 kilodaltons. Using cDNA probes, asteroid transcripts were localized to the proliferative tissues of embryos and to the mitotically active tissue anterior to the morphogenetic furrow in eye imaginal discs. Ribonuclease protection assays identified a mutation of asteroid that acts as a dominant enhancer of Star mutations and also enhances the Ellipse mutation, EgfrE1. Based on these data, a model for asteroid gene function in EGF receptor signaling is presented.Key words: Drosophila, asteroid, Star, EGF receptor, eye development.

scholarly journals Survey and Characterization of Viruses in Sweetpotato from Zimbabwe

Plant Disease ◽  
1997 ◽  
Vol 81 (10) ◽  
pp. 1115-1122 ◽  
Author(s):  
Farayi Chavi ◽  
A. Ian Robertson ◽  
Benedictus J. M. Verduin
Keyword(s):  
Amino Acids ◽  
Coat Protein ◽  
Sodium Dodecyl ◽  
Protein Band ◽  
Mottle Virus ◽  
Major Protein ◽  
Degenerate Primers ◽  
Vein Clearing ◽  
Major Protein Band

Thirty-one clones of sweetpotatoes collected from some parts of Zimbabwe were used as inoculum sources to mechanically inoculate 13 experimental hosts: Chenopodium amaranticolor, C. quinoa, Cucumis sativus, Datura stramoniumitalic, Gomphrena globosa, Ipomoea purpurea, I. quamoclit, I. rubrocorulea, Nicotiana benthamiana, N. clevelandii, N. glutinosa, N. rustica, and N. tabacum. Systemic vein clearing was observed in N. benthamiana inoculated with buffered sap from nine clones. Purification of the vein clearing inducing agent from one of the sweetpotato clones gave yields ranging from 2 to 17 mg/kg and the A260nm/A280nm was around 1.2. Electron microscopy revealed flexuous filamentous particles with a modal length of 830 nm. Protein analysis of purified virus preparations by sodium dodecyl sulfate polyacrylamide gel electrophoresis revealed a major protein band of 40 kDa, and this was assumed to be the viral coat protein. Minor protein bands of 27, 37, and 46 kDa were also observed. The viral protein degraded upon storage at 4°C over time to yield a protein band of 27 kDa. Polyclonal antiserum was produced against the purified virus. Protein A gold labeling of the purified virus incubated with available antisera; sweetpotato chlorotic stunt virus (SPCSV), sweetpotato feathery mottle virus strain russet crack (SPFMV-RC), sweetpotato feathery mottle virus, sweetpotato mild mottle virus, sweetpotato latent virus, sweetpotato chlorotic fleck virus, and sweetpotato caulimo-like virus resulted in a higher labeling density with the antiserum of SPFMV-RC than with the antiserum of SPCSV, while the other sera did not react. Further characterization of the vein clearing inducing agent was attempted by reverse transcription-polymerase chain reaction amplification of total RNA with degenerate primers for potyviruses and an oligo dT primer and PCR products of correct size were obtained. The nucleotide sequence was determined and the amino acid of the polyprotein deduced. Comparison with other strains of SPFMV showed strong similarity except for an insertion of 22 amino acids at the N-terminus of the coat protein. The coat protein size of 335 amino acids is the biggest SPFMV so far determined.

The investigation of nematocidal activity in Stenotrophomonas maltophilia G2 and characterization of a novel virulence serine protease

2009 ◽  
Vol 55 (8) ◽  
pp. 934-942 ◽  
Author(s):  
Xiaowei Huang ◽  
Junwei Liu ◽  
Junmei Ding ◽  
Qiusheng He ◽  
Rui Xiong ◽  
...  
Keyword(s):  
Serine Protease ◽  
Serine Proteases ◽  
Stenotrophomonas Maltophilia ◽  
Bursaphelenchus Xylophilus ◽  
Panagrellus Redivivus ◽  
Bacterial Culture Supernatant ◽  
Free Living Nematode ◽  
Nematocidal Activity ◽  
Nematode Cuticle

The Gram-negative bacterium Stenotrophomonas maltophilia G2 was isolated from a soil sample and was found to have high nematotoxic activity against a free-living nematode, Panagrellus redivivus, and a plant-parasitic nematode, Bursaphelenchus xylophilus . The analysis of virulence factors revealed that although the small molecular metabolites participated in nematode killing, the crude extracellular protein extract from the bacterial culture supernatant contributed significantly to its nematocidal activity. An extracellular protease was purified by chromatography, and its effects on degrading purified nematode cuticle and killing living nematodes were confirmed experimentally. Characterization of this purified protease revealed that the application of phenylmethylsulphonyl fluoride, an inhibitor of serine proteases, could completely abolish its proteolytic activity. The results from N-terminal amino acid sequencing showed no similarity with any known serine protease in S. maltophilia, suggesting a novel virulence serine protease was obtained. Our study is the first to show the nematocidal activity of S. maltophilia, and we identified a novel serine protease as an important pathogenicity factor.

scholarly journals Characterisation of chalcone isomerase gene isolated from Pueraria montana var.lobata plant

2021 ◽  
Vol 63 (12) ◽  
pp. 64-68
Author(s):  
Thi Bich Ngoc Tran ◽  
◽  
Tien Dung Nguyen ◽  
Thi Thu Hue Huynh ◽  
◽  
...  
Keyword(s):  
Amino Acids ◽  
Active Sites ◽  
Anthocyanin Biosynthesis ◽  
Alpha Helix ◽  
Amino Acid Sequences ◽  
Chalcone Isomerase ◽  
Bioinformatic Tools ◽  
Pueraria Montana ◽  
Complete Protein ◽  
Conserved Amino Acids

Chalcone isomerase (CHI) is well-known as an important enzyme in the biosynthetic pathways such as flavonoid, isoflavonoid, and anthocyanin biosynthesis. The enzyme was investigated in some kinds of plants in Fabaceae but no research was conducted about the CHI gene of Pueraria montana var. lobata (P. lobata) in Vietnam. In order to provide more information and characterisation of the gene, our study isolated the CHI gene by RT-PCR and Sangersequencing. The sequence of the CHIgene was analysed with nucleotide and deduced amino acid sequences to find the main domains. A full-length CDS of CHI gene from P. lobata is 672 bp encoded 224 amino acids. By using bioinformatic tools to compare, the isolated gene shared 99.7% homology with the same species reference (code D63577.1). Two different nucleotides in the gene were altered the amino acids in the protein, but the differences have not happened in active sites. Additionally, the conserved amino acids related to active catalysis of a hydrogen bond network also appeared in the P. lobataCHI gene. SWISS-MODEL was used to build the complete protein modeling showing that P. lobataCHI protein was the most similar with CHI of Medicago sativa - was defined structure in which all alpha-helix and beta-helix were completelyhomologies.

scholarly journals Characterization of mutants of three highly conserved amino acids in microsomal cytochrome P450 2B4

2006 ◽  
Vol 20 (4) ◽  
Author(s):  
Lucy Waskell ◽  
Haoming Zhang ◽  
Sang‐Choul Im ◽  
Roman Davydov ◽  
Brian Hoffman
Keyword(s):  
Amino Acids ◽  
Cytochrome P450 ◽  
Microsomal Cytochrome ◽  
Cytochrome P450 2B4 ◽  
Microsomal Cytochrome P450 ◽  
Conserved Amino Acids
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