scholarly journals Genetics of l-Sorbose Transport and Metabolism in Lactobacillus casei

2000 ◽  
Vol 182 (1) ◽  
pp. 155-163 ◽  
Author(s):  
María J. Yebra ◽  
Ana Veyrat ◽  
Mario A. Santos ◽  
Gaspar Pérez-Martínez
Keyword(s):  
Amino Acid ◽  
Catabolite Repression ◽  
Lactobacillus Casei ◽  
Amino Acid Sequences ◽  
Peptide Sequence ◽  
Transcriptional Analysis ◽  
Transcriptional Unit ◽  
Sugar Uptake ◽  
High Similarity ◽  
Chromosomal Dna

ABSTRACT Genes encoding l-sorbose metabolism ofLactobacillus casei ATCC 393 have been identified on a 6.8-kb chromosomal DNA fragment. Sequence analysis revealed seven complete genes and a partial open reading frame transcribed as two units. The deduced amino acid sequences of the first transcriptional unit (sorRE) showed high similarity to the transcriptional regulator and the l-sorbose-1-phosphate reductase of the sorbose (sor) operon from Klebsiella pneumoniae. The other genes are transcribed as one unit (sorFABCDG) in opposite direction to sorRE. The deduced peptide sequence of sorF showed homology with thed-sorbitol-6-phosphate dehydrogenase encoded in thesor operon from K. pneumoniae andsorABCD to components of the mannose phosphotransferase system (PTS) family but especially to domains EIIA, EIIB, EIIC and EIID of the phosphoenolpyruvate-dependentl-sorbose PTS from K. pneumoniae. Finally, the deduced amino acid sequence of a truncated gene (sorG) located downstream of sorD presented high similarity with ketose-1,6-bisphosphate aldolases. Results of studies on enzyme activities and transcriptional analysis revealed that the two gene clusters, sorRE and sorFABCDG, are induced by l-sorbose and subject to catabolite repression by d-glucose. Data indicating that the catabolite repression is mediated by components of the PTS elements and by CcpA, are presented. Results of sugar uptake assays inL. casei wild-type and sorBC mutant strains indicated that l-sorbose is taken up byl-sorbose-specific enzyme II and that L. caseicontains an inducible d-fructose-specific PTS. Results of growth analysis of those strains and a man sorBC double mutant suggested that l-sorbose is probably also transported by the d-mannose PTS. We also present evidence, from studies on a sorR mutant, suggesting that thesorR gene encodes a positive regulator of the twosor operons. Sequence alignment of SorR, SorC (K. pneumoniae), and DeoR (Bacillus subtilis) revealed that they might constitute a new group of transcriptional regulators.

scholarly journals AmyA, an α-Amylase with β-Cyclodextrin-Forming Activity, and AmyB from the Thermoalkaliphilic Organism Anaerobranca gottschalkii: Two α-Amylases Adapted to Their Different Cellular Localizations†

2005 ◽  
Vol 71 (7) ◽  
pp. 3709-3715 ◽  
Author(s):  
Meike Ballschmiter ◽  
Martin Armbrecht ◽  
Krasimira Ivanova ◽  
Garabed Antranikian ◽  
Wolfgang Liebl
Keyword(s):  
Amino Acid ◽  
Half Life ◽  
Extracellular Enzyme ◽  
Amino Acid Sequences ◽  
Cyclodextrin Glycosyltransferase ◽  
High Similarity ◽  
Ph Optimum ◽  
Transglycosylation Activity ◽  
Optimum Ph ◽  
Ph Range

ABSTRACT Two α-amylase genes from the thermophilic alkaliphile Anaerobranca gottschalkii were cloned, and the corresponding enzymes, AmyA and AmyB, were investigated after purification of the recombinant proteins. Based on their amino acid sequences, AmyA is proposed to be a lipoprotein with extracellular localization and thus is exposed to the alkaline milieu, while AmyB apparently represents a cytoplasmic enzyme. The amino acid sequences of both enzymes bear high similarity to those of GHF13 proteins. The different cellular localizations of AmyA and AmyB are reflected in their physicochemical properties. The alkaline pH optimum (pH 8), as well as the broad pH range, of AmyA activity (more than 50% activity between pH 6 and pH 9.5) mirrors the conditions that are encountered by an extracellular enzyme exposed to the medium of A. gottschalkii, which grows between pH 6 and pH 10.5. AmyB, on the other hand, has a narrow pH range with a slightly acidic pH optimum at 6 to 6.5, which is presumably close to the pH in the cytoplasm. Also, the intracellular AmyB is less tolerant of high temperatures than the extracellular AmyA. While AmyA has a half-life of 48 h at 70°C, AmyB has a half-life of only about 10 min at that temperature, perhaps due to the lack of stabilizing constituents of the cytoplasm. AmyA and AmyB were very similar with respect to their substrate specificity profiles, clearly preferring amylose over amylopectin, pullulan, and glycogen. Both enzymes also hydrolyzed α-, β-, and γ-cyclodextrin. Very interestingly, AmyA, but not AmyB, displayed high transglycosylation activity on maltooligosaccharides and also had significant β-cyclodextrin glycosyltransferase (CGTase) activity. CGTase activity has not been reported for typical α-amylases before. The mechanism of cyclodextrin formation by AmyA is unknown.

Comparative analyses of Serratia spp. outer membrane porin proteins

1993 ◽  
Vol 39 (4) ◽  
pp. 442-447 ◽  
Author(s):  
Joanne Hutsul ◽  
Elizabeth Worobec ◽  
Tom R. Parr Jr. ◽  
Gerald W. Becker
Keyword(s):  
Amino Acid ◽  
Serratia Marcescens ◽  
Single Channel ◽  
Enteric Bacteria ◽  
Amino Acid Sequences ◽  
Chromosomal Dna ◽  
Terminal Amino Acid ◽  
Molecular Weight Range ◽  
Terminal Amino ◽  
Porin Proteins

Eight Serratia strains and several members of the Enterobacteriaceae family were used in immunoblot and Southern DNA hybridization experiments and probed with antibody and DNA probes specific for the 41-kDa Serratia marcescens porin, to determine the extent of homology between Gram-negative porins. Immunoblot analyses performed using porin-specific rabbit sera and cell envelope preparations from these strains revealed that all strains produced at least one cross-reactive protein in the 41-kDa molecular weight range. Chromosomal DNA from each of the same strains was used in Southern analyses, probed with a 20-base-length oligonucleotide probe deduced from the N-terminal amino acid sequence of the 41-kDa Serratia marcescens porin. The probe hybridized to DNA from all of the Serratia species and six of the nine other enteric bacteria. Putative porin proteins from all the Serratia species were subjected to N-terminal amino acid sequencing and porin functional analysis using the black lipid bilayer method. All amino acid sequences were identical, with one exception in which an asparagine was substituted for an aspartic acid in Serratia rubidaea. All porins had very similar porin function (single channel conductance ranging between 1.72 and 2.00 nS). The results from this study revealed that a strong conservation exists among the Serratia porins and those produced by other enteric bacteria.Key words: porins, Serratia marcescens, homology studies.

Functional discrimination of sea anemone neurotoxins using 3D-plotting

2009 ◽  
Vol 4 (1) ◽  
pp. 41-49
Author(s):  
Ludis Morales ◽  
Orlando Acevedo ◽  
María Martínez ◽  
Dmitry Gokhman ◽  
Carlos Corredor
Keyword(s):  
Amino Acid ◽  
Dimensional Space ◽  
Three Dimensional ◽  
Sea Anemone ◽  
Amino Acid Sequences ◽  
Peptide Sequence ◽  
Sea Anemones ◽  
Functional Relationships ◽  
Voltage Dependent ◽  
Three Dimensional Space

AbstractOne of the most important goals in structural biology is the identification of functional relationships among the structure of proteins and peptides. The purpose of this study was to (1) generate a model based on theoretical and computational considerations among amino acid sequences within select neurotoxin peptides, and (2) compare the relationship these values have to the various toxins tested. We employed isolated neurotoxins from sea anemones with established specific potential to act on voltage-dependent sodium and potassium channel activity as our model. Values were assigned to each amino acid in the peptide sequence of the neurotoxins tested using the Number of Lareo and Acevedo algorithm (NULA). Once the NULA number was obtained, it was then plotted using three dimensional space coordinates. The results of this study allow us to report, for the first time, that there is a different numerical and functional relationship between the sequences of amino acids from sea anemone neurotoxins, and the resulting numerical relationship for each peptide, or NULA number, has a unique location in three-dimensional space.

scholarly journals Molecular cloning and deduced amino acid sequences of the γ-subunits of rat and monkey NAD+-isocitrate dehydrogenases

1993 ◽  
Vol 295 (2) ◽  
pp. 347-350 ◽  
Author(s):  
B J Nichols ◽  
L Hall ◽  
A C F Perry ◽  
R M Denton
Keyword(s):  
Amino Acid ◽  
Sequence Data ◽  
Amino Acid Sequences ◽  
Peptide Sequence ◽  
Cdna Libraries ◽  
Gamma Subunit ◽  
Gamma Subunits ◽  
Testis Cdna ◽  
Rat Epididymis ◽  
High Level

A 600 bp cDNA fragment encoding part of the gamma-subunit of pig heart NAD(+)-isocitrate dehydrogenase (ICDH gamma) was amplified by PCR using redundant oligonucleotide primers based on partial peptide sequence data [Huang and Colman (1990) Biochemistry 29, 8266-8273]. This PCR fragment was then used as a probe to isolate clones encoding the complete mature forms of the gamma-subunit from rat epididymis and monkey testis cDNA libraries. Comparison of the deduced amino acid sequences of the rat and monkey subunits and the partial sequence of the pig heart enzyme revealed a remarkably high level of sequence identity. The relationship between the deduced amino acid sequences of the NAD(+)-ICDH gamma-subunits and those of nonmammalian NAD(+)- and NADP(+)-ICDH subunits is discussed.

scholarly journals Molecular cloning of cDNA encoding an unrecognized component of amyloid in Alzheimer disease

1993 ◽  
Vol 90 (23) ◽  
pp. 11282-11286 ◽  
Author(s):  
K Uéda ◽  
H Fukushima ◽  
E Masliah ◽  
Y Xia ◽  
A Iwai ◽  
...  
Keyword(s):  
Amino Acid ◽  
Alzheimer Disease ◽  
Amyloid Fibrils ◽  
Protein A ◽  
Microscopic Analysis ◽  
Amino Acid Sequences ◽  
Full Length ◽  
Peptide Sequence ◽  
Electron Microscopic ◽  
Hydrophobic Domain

A neuropathological hallmark of Alzheimer disease (AD) is a widespread amyloid deposition. We analyzed the entire amino acid sequences in an amyloid preparation and found, in addition to the major beta/A4-protein (A beta) fragment, two unknown peptides. We raised antibodies against synthetic peptides using subsequences of these peptides. These antibodies immunostained amyloid in neuritic and diffuse plaques as well as vascular amyloid. Electron microscopic analysis demonstrated that the immunostaining was localized on amyloid fibrils. We have isolated an apparently full-length cDNA encoding a 140-amino-acid protein within which two previously unreported amyloid sequences are encoded in tandem in the most hydrophobic domain. We tentatively named this 35-amino acid peptide NAC (non-A beta component of AD amyloid) and its precursor NACP. NAC is the second component, after A beta, identified chemically in the purified AD amyloid preparation. Secondary structure predictions indicate that the NAC peptide sequence has a strong tendency to form beta-structures consistent with its association with amyloid. NACP is detected as a M(r) 19,000 protein in the cytosolic fraction of brain homogenates and comigrates on immunoblots with NACP synthesized in Escherichia coli from NACP cDNA. NACP mRNA is expressed principally in brain but is also expressed in low concentrations in all tissues examined except in liver, suggesting its ubiquitous and brain-specific functions. The availability of the cDNA encoding full-length NACP should help to elucidate the mechanisms of amyloidosis in AD.

scholarly journals Molecular cloning and deduced amino acid sequences of the α- and β- subunits of mammalian NAD+-isocitrate dehydrogenase

1995 ◽  
Vol 310 (3) ◽  
pp. 917-922 ◽  
Author(s):  
B J Nichols ◽  
A C F Perry ◽  
L Hall ◽  
R M Denton
Keyword(s):  
Amino Acid ◽  
Amino Acid Sequence ◽  
Isocitrate Dehydrogenase ◽  
Sequence Data ◽  
Amino Acid Sequences ◽  
Beta Subunit ◽  
Peptide Sequence ◽  
Alpha Subunit ◽  
Cdna Clones ◽  
Oligonucleotide Primers

A 153 bp fragment of the cDNA encoding the beta-subunit of pig heart NAD(+)-isocitrate dehydrogenase (NAD(+)-ICDH) was specifically amplified by PCR, using redundant oligonucleotide primers based on partial peptide sequence data [Huang and Colman (1990) Biochemistry 29, 8266-8273]. This PCR fragment was then used as a probe to isolate cDNA clones encoding the complete mature form of the beta-subunit from a monkey testis cDNA library. Examination of the deduced amino acid sequence of the monkey subunit and the partial sequence of the pig heart enzyme revealed a high level of sequence conservation. In addition, 3 overlapping fragments of the cDNA for the alpha-subunit of monkey NAD(+)-ICDH were amplified using oligonucleotide primers derived from the cDNA sequence of a subunit of bovine NAD(+)-ICDH (EMBL accession no: U07980). These cDNA fragments allow deduction of the amino acid sequence of the alpha-subunit. Since the gamma-subunit of monkey NAD(+)-ICDH has already been cloned [Nichols, Hall, Perry and Denton (1993) Biochem. J. 295, 347-350], a deduced amino acid sequence is now available for all three subunits of mammalian NAD(+)-ICDH. Interrelationships between these subunits are discussed and they are compared with the two subunits of yeast NAD(+)-ICDH and Escherichia coli NADP(+)-ICDH.

scholarly journals Identification of the mob Genes of Plasmid pSC101 and Characterization of a Hybrid pSC101-R1162 System for Conjugal Mobilization

2000 ◽  
Vol 182 (17) ◽  
pp. 4875-4881 ◽  
Author(s):  
Richard Meyer
Keyword(s):  
Amino Acid ◽  
Functional Unit ◽  
Chimeric Protein ◽  
Amino Acid Sequences ◽  
Transcriptional Unit ◽  
Dna Cleaving ◽  
Dna Base ◽  
Plasmid Mobilization ◽  
Dna Processing

ABSTRACT Similarities in DNA base sequence indicate that pSC101 and R1162 encode related systems for conjugal mobilization, although these plasmids are otherwise very different. The mob region of pSC101 was cloned, and two genes that are required for transfer were identified. One gene, mobA, encodes a protein similar in amino acid sequence to the DNA processing domain of the R1162 MobA protein. The other gene, mobX, is within the same transcriptional unit as the pSC101 mobA and is located just downstream, at the same position occupied by mobB in R1162. Despite this, the MobB and MobX proteins do not appear to be closely related based on a comparison of their amino acid sequences. Complementation analysis indicated that neither of the pSC101 Mob proteins could substitute for, or be replaced by, their R1162 counterparts, nor were they active together at the R1162 origin of transfer (oriT). However, the full set of R1162 Mob proteins did recognize the pSC101 oriT. A hybrid system for mobilization, active at the R1162 oriT site, was constructed. This system consists of MobX and a chimeric protein made up of the DNA cleaving-ligating domain of the R1162 MobA protein joined to a fragment of pSC101 MobA. Previous results suggested that MobB and a region of MobA distinct from the DNA processing domain together formed a functional unit in transfer. The present results support this model because the chimeric MobA, although active on R1162oriT, requires the pSC101 protein MobX for efficient plasmid mobilization.

scholarly journals Molecular biology of the 2-haloacid halidohydrolase IVa from Pseudomonas cepacia MBA4

1992 ◽  
Vol 284 (1) ◽  
pp. 87-93 ◽  
Author(s):  
U Murdiyatmo ◽  
W Asmara ◽  
J S H Tsang ◽  
A J Baines ◽  
A T Bull ◽  
...  
Keyword(s):  
Amino Acid ◽  
Nucleotide Sequence ◽  
Structural Gene ◽  
Sequence Data ◽  
Structural Data ◽  
Amino Acid Sequences ◽  
Open Reading Frames ◽  
Pseudomonas Cepacia ◽  
Chromosomal Dna ◽  
High Level

The structural gene (hdl IVa) for the Pseudomonas cepacia MBA4 2-haloacid halidohydrolase IVa (Hdl IVa) was isolated on a 1.6 kb fragment of Ps. cepacia MBA4 chromosomal DNA. The recombinant halidohydrolase was expressed in Escherichia coli and Pseudomonas putida and the structural gene was subcloned on to the tac expression vector pBTac1. High-level expression from the tac promoter was seen to be temperature-dependent, a consequence of the nucleotide sequence adjacent to the fragment encoding the halidohydrolase. The nucleotide sequence of the fragment encoding the Hdl IVa was determined and analysed. Three ATG codons were identified in one of the open reading frames and the one corresponding to the start of the hdl IVa structural gene was determined by comparison of the predicted amino acid sequences with the experimentally determined N-terminal sequences of halidohydrolase IVa. The hdl IVa gene encoded a 231-amino acid-residue protein of M(r) 25,900. The sequence and predicted structural data are discussed and comparison is made with sequence data for other halidohydrolases.

Molecular characterization and transcriptional analysis of the female-enriched chondroitin proteoglycan 2 of Toxocara canis

2017 ◽  
Vol 92 (2) ◽  
pp. 154-160
Author(s):  
G.X. Ma ◽  
R.Q. Zhou ◽  
L. Hu ◽  
Y.L. Luo ◽  
Y.F. Luo ◽  
...  
Keyword(s):  
Amino Acid ◽  
Toxocara Canis ◽  
Amino Acid Sequences ◽  
Transcriptional Analysis ◽  
Parasitic Nematodes ◽  
Biological Macromolecules ◽  
Limited Information ◽  
Reading Frame ◽  
Functional Studies ◽  
Nematode Parasites

AbstractToxocara canis is an important but neglected zoonotic parasite, and is the causative agent of human toxocariasis. Chondroitin proteoglycans are biological macromolecules, widely distributed in extracellular matrices, with a great diversity of functions in mammals. However, there is limited information regarding chondroitin proteoglycans in nematode parasites. In the present study, a female-enriched chondroitin proteoglycan 2 gene of T. canis (Tc-cpg-2) was cloned and characterized. Quantitative real-time polymerase chain reaction (qRT-PCR) was employed to measure the transcription levels of Tc-cpg-2 among tissues of male and female adult worms. A 485-amino-acid (aa) polypeptide was predicted from a continuous 1458-nuleotide open reading frame and designated as TcCPG2, which contains a 21-aa signal peptide. Conserved domain searching indicated three chitin-binding peritrophin-A (CBM_14) domains in the amino acid sequence of TcCPG2. Multiple alignment with the inferred amino acid sequences of Caenorhabditis elegans and Ascaris suum showed that CBM_14 domains were well conserved among these species. Phylogenetic analysis suggested that TcCPG2 was closely related to the sequence of chondroitin proteoglycan 2 of A. suum. Interestingly, a high level of Tc-cpg-2 was detected in female germline tissues, particularly in the oviduct, suggesting potential roles of this gene in reproduction (e.g. oogenesis and embryogenesis) of adult T. canis. The functional roles of Tc-cpg-2 in reproduction and development in this parasite and related parasitic nematodes warrant further functional studies.

scholarly journals Isolation, properties and amino acid sequences of three neurotoxins from the venom of a sea snake, Aipysurus laevis

1976 ◽  
Vol 153 (1) ◽  
pp. 79-87 ◽  
Author(s):  
N Maeda ◽  
N Tamiya
Keyword(s):  
Molecular Weight ◽  
Amino Acid ◽  
Sodium Dodecyl ◽  
Gel Filtration ◽  
Disc Electrophoresis ◽  
Amino Acid Sequences ◽  
Peptide Sequence ◽  
Amino Acid Residues ◽  
Toxin B ◽  
Almost All

Aipysurus laevis venom was chromatographed on CM-cellulose and Bio-Rex 70 columns. Three neurotoxic components, toxins Aipysurus laevis a, b and c, were isolated. The toxins a, b and c corresponded to 22, 33 and 21% respectively of the proteins in the original venom, and accounted for almost all the lethal activity of the venom. The three toxins a, b and c were monodisperse on disc electrophoresis at pH4; toxins a and b moved at the same velocity and c a little faster. They were monodisperse also on sodium dodecyl sulphate-polyacrylamide-disc-gel electrophoresis, giving a molecular weight of 7600. The molecular weight of toxin b estimated by gel filtration was 7000. The amino acid sequence analyses of these toxins revealed that they consisted of 60 amino acid residues and that Aipysurus laevis b was [25-methionine, 28-arginine] Aipysurus laevis a. Aipysurus laevis c was [28-lysine] Aipysurus laevis a, the tryptic peptide sequence relying on homology. The LD50 values of these toxins for 20g mice were 0.076 μg/g body wt. They inhibited the acetylcholine-induced contracture but did not affect the CKl-induced contracture of the isolated muscle.

Sign in / Sign up

Export Citation Format

Share Document