scholarly journals Cloning and Expression of the Gene for the Na+-Coupled Serine Transporter from Escherichia coli and Characteristics of the Transporter

1998 ◽  
Vol 180 (24) ◽  
pp. 6749-6752 ◽  
Author(s):  
Wakano Ogawa ◽  
Young-Mog Kim ◽  
Tohru Mizushima ◽  
Tomofusa Tsuchiya
Keyword(s):  
Escherichia Coli ◽  
Amino Acid ◽  
Amino Acid Sequence ◽  
Copy Number ◽  
Cloning And Expression ◽  
Amino Acid Residues ◽  
Low Copy Number ◽  
Hydropathy Analysis

ABSTRACT We cloned a gene (sstT) for the Na+/serine symporter from the chromosome of Escherichia coli by using a low-copy-number vector and sequenced it. According to the deduced amino acid sequence, the transporter (SstT) consists of 414 amino acid residues. Hydropathy analysis suggested that the SstT protein possesses 9, instead of 12, hydrophobic domains.

scholarly journals Diarrhea-Inducing Activity of Avian Rotavirus NSP4 Glycoproteins, Which Differ Greatly from Mammalian Rotavirus NSP4 Glycoproteins in Deduced Amino Acid Sequence, in Suckling Mice

2002 ◽  
Vol 76 (11) ◽  
pp. 5829-5834 ◽  
Author(s):  
Yoshio Mori ◽  
Mohammed Ali Borgan ◽  
Naoto Ito ◽  
Makoto Sugiyama ◽  
Nobuyuki Minamoto
Keyword(s):  
Escherichia Coli ◽  
Amino Acid ◽  
Amino Acid Sequence ◽  
Amino Acid Sequences ◽  
Amino Acid Residues ◽  
Suckling Mice

ABSTRACT Avian rotavirus NSP4 glycoproteins expressed in Escherichia coli acted as enterotoxins in suckling mice, as did mammalian rotavirus NSP4 glycoproteins, despite great differences in the amino acid sequences. The enterotoxin domain of PO-13 NSP4 exists in amino acid residues 109 to 135, a region similar to that reported in SA11 NSP4.

scholarly journals Phenotypic, Biochemical, and Genetic Analysis of KPC-41, a KPC-3 Variant Conferring Resistance to Ceftazidime-Avibactam and Exhibiting Reduced Carbapenemase Activity

2019 ◽  
Vol 63 (12) ◽  
Author(s):  
Linda Mueller ◽  
Amandine Masseron ◽  
Guy Prod’Hom ◽  
Tatiana Galperine ◽  
Gilbert Greub ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Amino Acids ◽  
Amino Acid ◽  
Klebsiella Pneumoniae ◽  
Genetic Analysis ◽  
Amino Acid Sequence ◽  
Cloning And Expression ◽  
Content Type ◽  
Amino Acid Insertion

ABSTRACT A novel KPC variant, KPC-41, was identified in a Klebsiella pneumoniae clinical isolate from Switzerland. This β-lactamase possessed a 3-amino-acid insertion (Pro-Asn-Lys) located between amino acids 269 and 270 compared to the KPC-3 amino acid sequence. Cloning and expression of the blaKPC-41 gene in Escherichia coli, followed by determination of MIC values and kinetic parameters, showed that KPC-41, compared to those of KPC-3, has an increased affinity to ceftazidime and a decreased sensitivity to avibactam, leading to resistance to ceftazidime-avibactam once produced in K. pneumoniae. Furthermore, KPC-41 exhibited a drastic decrease of its carbapenemase activity. This report highlights that a diversity of KPC variants conferring resistance to ceftazidime-avibactam already circulate in Europe.

scholarly journals The overexpression and complete amino acid sequence of Escherichia coli 3-dehydroquinase

1986 ◽  
Vol 238 (2) ◽  
pp. 475-483 ◽  
Author(s):  
K Duncan ◽  
S Chaudhuri ◽  
M S Campbell ◽  
J R Coggins
Keyword(s):  
Escherichia Coli ◽  
Amino Acid ◽  
Nucleotide Sequence ◽  
Amino Acid Sequence ◽  
Polypeptide Chain ◽  
Amino Acid Residues ◽  
Transcript Mapping ◽  
Terminal Amino Acid ◽  
Terminal Amino ◽  
Terminal Amino Acid Sequence

The enzyme 3-dehydroquinase was purified in milligram quantities from an overproducing strain of Escherichia coli. The amino acid sequence was deduced from the nucleotide sequence of the aroD gene and confirmed by determining the amino acid composition of the overproduced enzyme and its N-terminal amino acid sequence. The complete polypeptide chain consists of 240 amino acid residues and has a calculated subunit Mr of 26,377. Transcript mapping revealed that aroD is a typical monocistronic gene.

scholarly journals The cloning and expression of the aroL gene from Escherichia coli K12. Purification and complete amino acid sequence of shikimate kinase II, the aroL-gene product

1986 ◽  
Vol 237 (2) ◽  
pp. 427-437 ◽  
Author(s):  
G Millar ◽  
A Lewendon ◽  
M G Hunter ◽  
J R Coggins
Keyword(s):  
Escherichia Coli ◽  
Amino Acid ◽  
Amino Acid Sequence ◽  
Transcriptional Start Site ◽  
Cloning And Expression ◽  
Potential Operator ◽  
Gene Encoding ◽  
Shikimate Kinase ◽  
Complete Amino Acid Sequence ◽  
Escherichia Coli K12

The aroL gene encoding the enzyme shikimate kinase II was cloned from Escherichia coli K12. Construction of over-expressing strains permitted for the first time the purification to homogeneity of a monofunctional shikimate kinase. The complete amino acid sequence of shikimate kinase II was determined by a combined nucleotide and direct amino acid sequencing strategy. E. coli shikimate kinase II is a monomeric enzyme containing 173 amino acid residues with a calculated Mr 18,937. The amino acid sequence contains a region homologous with other kinases and ATP-requiring enzymes. Evidence is presented suggesting that the transcriptional start site of the aroL gene is located within a potential operator site.

scholarly journals Cloning and Expression of Islandisin, a New Thermostable Subtilisin from Fervidobacterium islandicum, in Escherichia coli

2005 ◽  
Vol 71 (7) ◽  
pp. 3951-3958 ◽  
Author(s):  
Carolin Gödde ◽  
Kerstin Sahm ◽  
Stan J. J. Brouns ◽  
Leon D. Kluskens ◽  
John van der Oost ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Amino Acid ◽  
Amino Acid Sequence ◽  
Serine Protease ◽  
Signal Sequence ◽  
Thermophilic Bacterium ◽  
Cloning And Expression ◽  
Inverse Pcr ◽  
Heat Denaturation ◽  
Catalytic Residues

ABSTRACT A gene encoding a subtilisin-like protease, designated islandisin, from the extremely thermophilic bacterium Fervidobacterium islandicum (DSMZ 5733) was cloned and actively expressed in Escherichia coli. The gene was identified by PCR using degenerated primers based on conserved regions around two of the three catalytic residues (Asp, His, and Ser) of subtilisin-like serine protease-encoding genes. Using inverse PCR regions flanking the catalytic residues, the gene could be cloned. Sequencing revealed an open reading frame of 2,106 bp. The deduced amino acid sequence indicated that the enzyme is synthesized as a proenzyme with a putative signal sequence of 33 amino acids (aa) in length. The mature protein contains the three catalytic residues (Asp177, His215, and Ser391) and has a length of 668 aa. Amino acid sequence comparison and phylogenetic analysis indicated that this enzyme could be classified as a subtilisin-like serine protease in the subgroup of thermitase. The whole gene was amplified by PCR, ligated into pET-15b, and successfully expressed in E. coli BL21(DE3)pLysS. The recombinant islandisin was purified by heat denaturation, followed by hydroxyapatite chromatography. The enzyme is active at a broad range of temperatures (60 to 80°C) and pHs (pH 6 to 8.5) and shows optimal proteolytic activity at 80°C and pH 8.0. Islandisin is resistant to a number of detergents and solvents and shows high thermostability over a long period of time (up to 32 h) at 80°C with a half-life of 4 h at 90°C and 1.5 h at 100°C.

scholarly journals The overexpression, purification and complete amino acid sequence of chorismate synthase from Escherichia coli K12 and its comparison with the enzyme from Neurospora crassa

1988 ◽  
Vol 251 (2) ◽  
pp. 313-322 ◽  
Author(s):  
P J White ◽  
G Millar ◽  
J R Coggins
Keyword(s):  
Escherichia Coli ◽  
Amino Acid ◽  
Amino Acid Sequence ◽  
Neurospora Crassa ◽  
Gel Filtration ◽  
Cross Linking ◽  
Amino Acid Residues ◽  
Chorismate Synthase ◽  
E Coli ◽  
A Subunit

The enzyme chorismate synthase was purified in milligram quantities from an overproducing strain of Escherichia coli. The amino acid sequence was deduced from the nucleotide sequence of the aroC gene and confirmed by determining the N-terminal amino acid sequence of the purified enzyme. The complete polypeptide chain consists of 357 amino acid residues and has a calculated subunit Mr of 38,183. Cross-linking and gel-filtration experiments show that the enzyme is tetrameric. An improved purification of chorismate synthase from Neurospora crassa is also described. Cross-linking and gel-filtration experiments on the N. crassa enzyme show that it is also tetrameric with a subunit Mr of 50,000. It is proposed that the subunits of the N. crassa enzyme are larger because they contain a diaphorase domain that is absent from the E. coli enzyme.

scholarly journals Cloning and expression of diadenosine 5′,5‴-P1,P4-tetraphosphate hydrolase from Lupinus angustifolius L

1998 ◽  
Vol 329 (2) ◽  
pp. 313-319 ◽  
Author(s):  
Danuta MAKSEL ◽  
Andrzej GURANOWSKI ◽  
C. Steven ILGOUTZ ◽  
Arthur MOIR ◽  
G. Michael BLACKBURN ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Amino Acids ◽  
Amino Acid ◽  
Amino Acid Sequence ◽  
Sequence Similarity ◽  
Lupinus Angustifolius ◽  
Cloning And Expression ◽  
Lupinus Luteus ◽  
Higher Plant ◽  
Polyadenylated Rna

The first isolation, cloning and expression of cDNA encoding an asymmetric diadenosine 5ʹ,5‴P1,P4-tetraphosphate pyrophosphohydrolase (Ap4A hydrolase) from a higher plant is described. Ap4A hydrolase protein was purified from seeds of both Lupinus luteus and Lupinus angustifolius and partially sequenced. The Ap4A hydrolase cDNA was cloned from L. angustifolius cotyledonary polyadenylated RNA using reverse transcription and PCR with primers based on the amino acid sequence. The cDNA encoded a protein of 199 amino acids, molecular mass 22982 Da. When expressed in Escherichia coli fused to a maltose-binding protein, the enzyme catalysed asymmetric cleavage of Ap4A to AMP and ATP which was inhibited at concentrations of F- as low as 3 μM. These are properties characteristic of Ap4A hydrolase (asymmetrical) (EC 3.6.1.17). Comparison of the Ap4A hydrolase sequences derived from the four known cDNAs from pig, human, lupin and fission yeast showed that, like the mammalian hydrolase, the lupin enzyme possesses a Mut T motif but no other significant similarities. No sequence similarity to the human fragile histidine triad protein, as found in the Ap4A hydrolase from Schizosaccharomyces pombe, was detected in the Ap4A hydrolase from lupin.

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