scholarly journals Characterization of a Thermostable l-Arabinose (d-Galactose) Isomerase from the Hyperthermophilic Eubacterium Thermotoga maritima

2004 ◽  
Vol 70 (3) ◽  
pp. 1397-1404 ◽  
Author(s):  
Dong-Woo Lee ◽  
Hyeung-Jin Jang ◽  
Eun-Ah Choe ◽  
Byoung-Chan Kim ◽  
Sang-Jae Lee ◽  
...  
Keyword(s):  
Amino Acid ◽  
Elevated Temperatures ◽  
Thermotoga Maritima ◽  
Sequence Similarity ◽  
Gel Filtration ◽  
Catalytic Efficiency ◽  
Maximal Activity ◽  
Recombinant Enzyme ◽  
Amino Acid Sequence Similarity ◽  
Calculated Molecular Weight

ABSTRACT The araA gene encoding l-arabinose isomerase (AI) from the hyperthermophilic bacterium Thermotoga maritima was cloned and overexpressed in Escherichia coli as a fusion protein containing a C-terminal hexahistidine sequence. This gene encodes a 497-amino-acid protein with a calculated molecular weight of 56,658. The recombinant enzyme was purified to homogeneity by heat precipitation followed by Ni2+ affinity chromatography. The native enzyme was estimated by gel filtration chromatography to be a homotetramer with a molecular mass of 232 kDa. The purified recombinant enzyme had an isoelectric point of 5.7 and exhibited maximal activity at 90�C and pH 7.5 under the assay conditions used. Its apparent Km values for l-arabinose and d-galactose were 31 and 60 mM, respectively; the apparent V max values (at 90�C) were 41.3 U/mg (l-arabinose) and 8.9 U/mg (d-galactose), and the catalytic efficiencies (k cat/Km ) of the enzyme were 74.8 mM−1 � min−1 (l-arabinose) and 8.5 mM−1 � min−1 (d-galactose). Although the T. maritima AI exhibited high levels of amino acid sequence similarity (>70%) to other heat-labile mesophilic AIs, it had greater thermostability and higher catalytic efficiency than its mesophilic counterparts at elevated temperatures. In addition, it was more thermostable in the presence of Mn2+ and/or Co2+ than in the absence of these ions. The enzyme carried out the isomerization of d-galactose to d-tagatose with a conversion yield of 56% for 6 h at 80�C.

scholarly journals Identification and Characterization of a Novel Intracellular Alkaline α-Amylase from the Hyperthermophilic Bacterium Thermotoga maritima MSB8

2006 ◽  
Vol 72 (3) ◽  
pp. 2206-2211 ◽  
Author(s):  
Meike Ballschmiter ◽  
Ole Fütterer ◽  
Wolfgang Liebl
Keyword(s):  
Amino Acid ◽  
Amino Acid Sequence ◽  
Thermotoga Maritima ◽  
Sequence Similarity ◽  
Hydrolytic Activity ◽  
Amino Acid Sequence Similarity ◽  
Glycoside Hydrolase Family ◽  
Ph Optimum ◽  
Intracellular Enzyme ◽  
Hyperthermophilic Bacterium

ABSTRACT The gene for a novel α-amylase, designated AmyC, from the hyperthermophilic bacterium Thermotoga maritima was cloned and heterologously overexpressed in Escherichia coli. The putative intracellular enzyme had no amino acid sequence similarity to glycoside hydrolase family (GHF) 13 α-amylases, yet the range of substrate hydrolysis and the product profile clearly define the protein as an α-amylase. Based on sequence similarity AmyC belongs to a subgroup within GHF 57. On the basis of amino acid sequence similarity, Glu185 and Asp349 could be identified as the catalytic residues of AmyC. Using a 60-min assay, the maximum hydrolytic activity of the purified enzyme, which was dithiothreitol dependent, was found to be at 90°C. AmyC displayed a remarkably high pH optimum of pH 8.5 and an unusual sensitivity towards both ATP and EDTA.

scholarly journals Site-directed mutagenesis at aspartate and glutamate residues of xylanase from Bacillus pumilus

1992 ◽  
Vol 288 (1) ◽  
pp. 117-121 ◽  
Author(s):  
E P Ko ◽  
H Akatsuka ◽  
H Moriyama ◽  
A Shinmyo ◽  
Y Hata ◽  
...  
Keyword(s):  
Amino Acids ◽  
Reaction Mechanism ◽  
Amino Acid ◽  
Bacillus Pumilus ◽  
Specific Activity ◽  
Sequence Similarity ◽  
Site Directed Mutagenesis ◽  
Dimensional Structure ◽  
Amino Acid Sequence Similarity ◽  
Directed Mutagenesis

To elucidate the reaction mechanism of xylanase, the identification of amino acids essential for its catalysis is of importance. Studies have indicated the possibility that the reaction mechanism of xylanase is similar to that of hen's egg lysozyme, which involves acidic amino acid residues. On the basis of this assumption, together with the three-dimensional structure of Bacillus pumilus xylanase and its amino acid sequence similarity to other xylanases of different origins, three acidic amino acids, namely Asp-21, Glu-93 and Glu-182, were selected for site-directed mutagenesis. The Asp residue was altered to either Ser or Glu, and the Glu residues to Ser or Asp. The purified mutant xylanases D21E, D21S, E93D, E93S, E182D and E182S showed single protein bands of about 26 kDa on SDS/PAGE. C.d. spectra of these mutant enzymes show no effect on the secondary structure of xylanase, except that of D21E, which shows a little variation. Furthermore, mutations of Glu-93 and Glu-182 resulted in a drastic decrease in the specific activity of xylanase as compared with mutation of Asp-21. On the basis of these results we propose that Glu-93 and Glu-182 are the best candidates for the essential catalytic residues of xylanase.

Na(+)-independent transport (uniport) of amino acids and glucose in mammalian cells.

1994 ◽  
Vol 196 (1) ◽  
pp. 93-108
Author(s):  
D K Kakuda ◽  
C L MacLeod
Keyword(s):  
Amino Acids ◽  
Amino Acid ◽  
Mammalian Cells ◽  
Glucose Transporter ◽  
Sequence Similarity ◽  
Amino Acid Sequence Similarity ◽  
Amino Acid Transporters ◽  
Cationic Amino Acid ◽  
Transporter Family ◽  
Share Amino Acid Sequence

Recent advances have made possible the isolation of the genes and their cDNAs encoding Na(+)-independent amino acid transporters. Two classes of amino acid 'uniporters' have been isolated. One class contains the mCAT (murine cationic amino acid transporter) gene family that encodes proteins predicted to span the membrane 12-14 times and exhibits structural properties similar to the GLUT (glucose transporter) family and to other well-known transporters. The other class consists of two known genes, rBAT (related to B system amino acid transporters) and 4F2hc, that share amino acid sequence similarity with alpha-amylases and alpha-glucosidases. They are type II glycoproteins predicted to span the membrane only once, yet they mediate the Na(+)-independent transport of cationic and zwitterionic amino acids in Xenopus oocytes. Mutations in the human rBAT gene have been identified by Palacín and his co-workers in several families suffering from a heritable form of cystinuria. This important finding clearly establishes a key role for rBAT in cystine transport. The two classes of amino acid transporters are compared with the well-studied GLUT family of Na(+)-independent glucose transporters.

History, Diagnosis, and Treatment of Coronavirus Disease 2019 (COVID-19)

Coronaviruses ◽  
2021 ◽  
Vol 02 ◽  
Author(s):  
Amaresh Mishra ◽  
Nisha Nair ◽  
Vishwas Tripathi ◽  
Yamini Pathak ◽  
Jaseela Majeed
Keyword(s):  
Amino Acid ◽  
Severe Acute Respiratory Syndrome ◽  
Sequence Similarity ◽  
Neurological Complications ◽  
Amino Acid Sequences ◽  
World Health ◽  
Amino Acid Sequence Similarity ◽  
Effective Prevention ◽  
Short Period ◽  
The World

: The Coronavirus Disease 2019 (COVID-19), also known as a novel coronavirus (2019-nCoV), reportedly originated from Wuhan City, Hubei Province, China. Coronavirus Disease 2019 rapidly spread all over the world within a short period. On January 30th, 2020, the World Health Organization (WHO) declared it a global epidemic. COVID-19 is a severe acute respiratory syndrome coronavirus (SARS-CoV) virus that evolves to respiratory, hepatic, gastrointestinal, and neurological complications, and eventually death. SARS-CoV and the Middle East Respiratory Syndrome coronavirus (MERS-CoV) genome sequences similar identity with 2019-nCoV or severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). However, few amino acid sequences of 2019-nCoV differ from SARS-CoV and MERS-CoV. COVID-19 shares about 90% amino acid sequence similarity with SARS-CoV. Effective prevention methods should be taken in order to control this pandemic situation. Till now, there are no effective treatments available to treat COVID-19. This review provides information regarding COVID-19 history, epidemiology, pathogenesis, and molecular diagnosis. Also, we focus on the development of vaccines in the management of this COVID-19 pandemic and limiting the spread of the virus.

The gene encoding a major component of the lateral elements of synaptonemal complexes of the rat is related to X-linked lymphocyte-regulated genes

1994 ◽  
Vol 14 (2) ◽  
pp. 1137-1146
Author(s):  
J H Lammers ◽  
H H Offenberg ◽  
M van Aalderen ◽  
A C Vink ◽  
A J Dietrich ◽  
...  
Keyword(s):  
Amino Acid ◽  
Sequence Similarity ◽  
Sodium Dodecyl ◽  
Coiled Coil ◽  
Amino Acid Identity ◽  
Male Rat ◽  
Amino Acid Sequence Similarity ◽  
Gene Encoding ◽  
Synaptonemal Complexes ◽  
Complex Protein

The lateral elements of synaptonemal complexes (SCs) of the rat contain major components with relative electrophoretic mobilities (M(r)S) of 30,000 and 33,000. After one-dimensional separation of SC proteins on polyacrylamide-sodium dodecyl sulfate gels, these components show up as two broad bands. These bands contain closely related proteins, as judged from their peptide maps and immunological reactivity. Using affinity-purified polyclonal anti-30,000- and anti-33,000-M(r) component antibodies, we isolated a cDNA encoding at least one of the 30,000- or 33,000-M(r) SC components. The protein predicted from the nucleotide sequence of the cDNA, called SCP3 (for synaptonemal complex protein 3), has a molecular mass of 29.7 kDa and a pI value of 9.4. It has a potential nucleotide binding site and contains stretches that are predicted to be capable of forming coiled-coil structures. In the male rat, the gene encoding SCP3 is transcribed exclusively in the testis. SCP3 has significant amino acid similarity to the pM1 protein, which is one of the predicted products of an X-linked lymphocyte-regulated gene family of the mouse: there are 63% amino acid sequence similarity and 35% amino acid identity between the SCP3 and pM1 proteins. However, SCP3 differs from pM1 in several respects, and whether the proteins fulfill related functions is still an open question.

scholarly journals Engineered Resistance Against Papaya ringspot virus in Venezuelan Transgenic Papayas

Plant Disease ◽  
2004 ◽  
Vol 88 (5) ◽  
pp. 516-522 ◽  
Author(s):  
Gustavo Fermin ◽  
Valentina Inglessis ◽  
Cesar Garboza ◽  
Sairo Rangel ◽  
Manuel Dagert ◽  
...  
Keyword(s):  
Amino Acid ◽  
Agrobacterium Tumefaciens ◽  
Coat Protein ◽  
Amino Acid Sequence ◽  
Sequence Similarity ◽  
Ringspot Virus ◽  
Amino Acid Sequence Similarity ◽  
Papaya Ringspot Virus ◽  
Local Varieties ◽  
Cp Gene

Local varieties of papaya grown in the Andean foothills of Mérida, Venezuela, were transformed independently with the coat protein (CP) gene from two different geographical Papaya ringspot virus (PRSV) isolates, designated VE and LA, via Agrobacterium tumefaciens. The CP genes of both PRSV isolates show 92 and 96% nucleotide and amino acid sequence similarity, respectively. Four PRSV-resistant R0 plants were intercrossed or selfed, and the progenies were tested for resistance against the homologous isolates VE and LA, and the heterologous isolates HA (Hawaii) and TH (Thailand) in greenhouse conditions. Resistance was affected by sequence similarity between the transgenes and the challenge viruses: resistance values were higher for plants challenged with the homologous isolates (92 to 100% similarity) than with the Hawaiian (94% similarity) and, lastly, Thailand isolates (88 to 89% similarity). Our results show that PRSV CP gene effectively protects local varieties of papaya against homologous and heterologous isolates of PRSV.

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