Similar Sequence Identity Research Articles

Measuring Homologous Recombination Rates between Chromosomal Locations in Salmonella.

Homologous recombination between two similar DNA molecules, plays an important role in the repair of double-stranded DNA breaks. Recombination can occur between two sister chromosomes, or between two locations of similar sequence identity within the same chromosome. The assay described here is designed to measure the rate of homologous recombination between two locations with sequence similarity within the same bacterial chromosome. For this purpose, a selectable/counter-selectable genetic cassette is inserted into one of the locations and homologous recombination repair rates are measured as a function of recombinational removal of the inserted cassette. This recombinational repair process is called gene conversion, non-reciprocal recombination. We used this method to measure the recombination rates between genes within gene families and to study the stability of mobile genetic elements inserted into members of gene families.

Effect of biofilm forming plant growth promoting rhizobacteria on salinity tolerance in barley

Formation of biofilm under varying stress conditions is a significant strategy adopted by bacterial strains for their successful survival in plant rhizosphere. In this study, the activity of biofilm formation of 20 isolates and strains of plant growth promoting rhizobacteria (PGPR) was determined under different salt concentrations. The results indicated that all of the 20 PGPRs have the activity of biofilm formation under 0.0, 250, 500 or 1000mM NaCl which was increased with increasing salt concentration. PGPR strains with the highest activity of biofilm formation were selected and used to coat barley grains. The coated grains were sown in clay/sandy soil and left to grow for 25 days. The results showed that bacterial inoculation was effective in alleviating the deleterious effect of salinity on some growth criteria (seedling length, fresh and dry masses as well as relative water content), compared with the control. The isolate HM6 (B6), which showed the highest activity of biofilm formation at all the studied NaCl concentrations, was identified using 16S ribosomal RNA gene amplification and sequencing of the PCR product. The similarity sequence analysis indicated that HM6 isolate has 97.4% similar sequence identity to Bacillus amyloliquifaciens. It could be speculated that the bacterial activity of biofilm formation is helpful for improving salt stress tolerance of barley.

Structural insight into the evolution of a new chemokine family from zebrafish

The mammalian chemokine family is segregated into four families - CC, CXC, CX3C, and XC-based on the arrangement of cysteines and the corresponding disulfides. Sequencing of the Danio rerio (zebrafish) genome has identified more than double the amount of human chemokines with the absence of the CX3C family and the presence of a new family, CX. The only other family with a single cysteine in the N-terminal region is the XC family. Human lymphotactin (XCL1) has two interconverting structures due to dynamic changes that occur in the protein. Similar to an experiment with XCL1 that identified the two structural forms, we probed for multiple forms of zCXL1 using heparin affinity. The results suggest only a single form of CXL1 is present. We used sulfur-SAD phasing to determine the three-dimensional structure CXL1. Zebrafish CXL1 (zCXL1) has three disulfides that appear to be important for a stable structure. One disulfide is common to all chemokines except those that belong to the XC family, another is similar to a subset of CC chemokines containing three disulfides, but the third disulfide is unique to the CX family. We analyzed the electrostatic potential of the zCXL1 structure and identified the likely heparin-binding site for glycosaminoglycans (GAGs). zCXL1 has a similar sequence identity with human CCL5 and CXCL12, but the structure is more related to CCL5. Our structural analysis supports the phylogenetic and genomic studies on the evolution of the CXL family.

Phylogenetic and DNA methylation analysis reveal novel regions of variable methylation in the mouse IAP class of transposons

BackgroundSelect retrotransposons in the long terminal repeat (LTR) class exhibit interindividual variation in DNA methylation that is altered by developmental environmental exposures. Yet, neither the full extent of variability at these “metastable epialleles,” nor the phylogenetic relationship underlying variable elements is well understood. The murine metastable epialleles, Avy and CabpIAP, result from independent insertions of an intracisternal A particle (IAP) mobile element, and exhibit remarkably similar sequence identity (98.5%).ResultsUtilizing the C57BL/6 genome we identified 10802 IAP LTRs overall and a subset of 1388 in a family that includes Avy and CabpIAP. Phylogenetic analysis revealed two duplication and divergence events subdividing this family into three clades. To characterize interindividual variation across clades, liver DNA from 17 isogenic mice was subjected to combined bisulfite and restriction analysis (CoBRA) for 21 separate LTR transposons (7 per clade). The lowest and highest mean methylation values were 59% and 88% respectively, while methylation levels at individual LTRs varied widely, ranging from 9% to 34%. The clade with the most conserved elements had significantly higher mean methylation across LTRs than either of the two diverged clades (p = 0.040 and p = 0.017). Within each mouse, average methylation across all LTRs was not significantly different (71%-74%, p > 0.99).ConclusionsCombined phylogenetic and DNA methylation analysis allows for the identification of novel regions of variable methylation. This approach increases the number of known metastable epialleles in the mouse, which can serve as biomarkers for environmental modifications to the epigenome.

Identification of CBL Genes from Foxtail Millet ( Setaria italica [L.] Beauv. ) and Its Expression under Drought and Salt Stresses

CBL/CIPK signal network system plays an important role in plant stress response. In this study, we identified seven candidate CBL genes designated as SiCBL1–7 from foxtail millet (Setaria italica [L.] Beauv.) genome using bioinformatics methods. The distributions of the predicted foxtail millet CBL genes were uneven in the nine chromosomes. Sequence analysis showed that the protein sequences and structure of foxtail millet CBL genes were very conservative. All the putative CBL genes contained 7–8 introns. Most exons of those SiCBL genes contained the same base number and shared highly similar sequence identity. All foxtail millet CBL genes consisted of four EF-Hand functional domains and the distance between the EF-hand motifs was conservative. Evolutionary analysis revealed that the CBL genes existed in the early evolutionary stage of terrestrial plants and all CBL genes were divided into four subgroups. The expression patterns of those SiCBL genes under drought and salt stresses, were displayed by RT-PCR and the results showed that four SiCBL genes (SiCBL1, SiCBL2, SiCBL3, and SiCBL5) were strongly induced by drought and three SiCBL genes (SiCBL1, SiCBL3, and SiCBL7) were induced by salt stress. SiCBL3 maybe takes an important role in foxtail millet under drought because of its constitutive high level expression. Foxtail millet CBL genes reported in this study would enrich CBL members in plant kingdom and lay a foundation for studying the function and mechanisms of the CBL/CIPK network system response to stresses.

Crystallization and preliminary X-ray diffraction studies of the abscisic acid receptor PYL3 and its complex with pyrabactin.

Abscisic acid (ABA) modulates many developmental processes and responses to environmental stress. Recently, a family of pyrabactin resistance-like proteins (PYLs) in Arabidopsis thaliana were identified to be abscisic acid receptors. Although the 14 PYLs members share a similar sequence identity, they exhibit different responses toward pyrabactin. Apo-PYL3 is a dimer; however, its oligomeric state changes greatly on the addition of pyrabactin. Moreover, pyrabactin binds dimeric PYL3 in a nonproductive mode which prevents receptor activation and inhibition of PP2Cs. Here, the expression, purification and crystallization of apo-PYL3 and of PYL3 complexed with pyrabactin are reported. Diffraction data were optimized to 2.5 Å resolution for apo-PYL3 and to 1.83 Å resolution for PYL3-pyrabactin. The crystals of apo-PYL3 and PYL3-pyrabactin belonged to space groups P4(1)2(1)2 and P2(1)2(1)2(1), respectively.

Sequence Comparison of Mitochondrial 16s rRNA Gene Segment in Penaeids

A partial fragment of 16s rRNA mitochondrial gene was amplified by PCR from Penaeus monodon samples collected from the west and east coast regions of India. The amplified 520 bp of 16s rRNA mitochondrial gene segment of P. monodon from different locations on sequencing revealed similar sequence identity and no apparent sequence variation could be observed in this amplified segment. Amplification of 16s rRNA mitochondrial gene segment was also carried out from Marsupenaeus japonicus and Fenneropenaeus indicus. The sequence identity of 16s rRNA mitochondrial gene segments between P. monodon and M. japonicus, between P. monodon and F. indicus and between M. japonicus and F. indicus was found to be 91%, 92% and 91% respectively. The high level of sequence identity observed in the amplified PCR products of 16s rRNA mitochondrial gene segment of P. monodon isolated from the east and west coast regions of India, suggests that it will be more useful to focus on other variable mitochondrial DNA regions which may be more informative for population studies.

Genetic diversity of epidemic enterovirus 71 strains recovered from clinical and environmental samples in Taiwan

Most enteroviruses excreted in human feces and urine are present in environmental water. In order to clarify the infection route of enterovirus, the detection of viruses in both clinical and environmental samples may contribute to understanding the mode of transmission of strains responsible for human infection. Thus, 21 epidemic enterovirus 71 strains from environmental water or stool samples were collected from HFMD children during 2005. Enterovirus genomic RNA was first amplified directly from clinical and environmental samples and then characterized by DNA sequencing and phylogenetic analysis. Results showed that these clinical strains share similar sequence identity (86.4–86.8%) to prototype BrCr based on the 5′-nontranslated region (NTR). However, environmental strains, except HME 77, share similar sequence identity (86.2–87.2%) to prototype BrCr. HME 77 showed higher sequence identity (90.1%). Results from phylogenetic analysis revealed that five environmental isolates were clustered as genogroup 3, which was closely related to a Taiwan outbreak in 1998 ( AY055133). HME 77 was more closely related to a China epidemic isolate ( AY895144), which belonged to genogroup 4. In contrast, all strains from clinical samples tested belonged to genogroup 3, which clustered with AY055133. In conclusion, there are two major epidemic clones (genogroups 3 and 4) prevalent in Taiwan since 2004 either in water or clinical patients.

A pepD-like peptidase from the ruminal bacterium, Prevotella albensis

Peptidases of Prevotella spp. play an important role in the breakdown of protein to ammonia in the rumen. This study describes a peptidase cloned from Prevotella albensis M384. DNA from P. albensis was used to complement a peptidase-deficient strain of Escherichia coli, CM107. A cloned fragment, Pep581, which enabled growth of E. coli CM107, contained an ORF of 1452 bp, encoding a 484 amino acid residue protein with a calculated molecular weight of 53.2 kDa and a theoretical pI of 4.90. Pep581 shared similar sequence identity of 47% with PepD from E. coli, and it was also a metallo-aminopeptidase. A putative catalytic metal binding region was identified in Pep581, similar to that found in the related PepT (a tripeptidase) and PepA (an oligopeptidase). Gel filtration indicated Pep581 was a dimer in its native state, similar to PepD of E. coli. PepD is a broad specificity dipeptidase that has been found in several prokaryotes. The enzyme expressed from Pep581 differed from PepD enzymes previously characterised in that it hydrolysed tri- and oligopeptides in addition to dipeptides, cleaving single amino acids from the N terminus.

A new natriuretic peptide isolated from cardiac atria of trout, Oncorhynchus mykiss

Atrial and brain natriuretic peptides (ANP and BNP, respectively) are two cardiac natriuretic peptides (NPs) found in tetrapods from amphibians to mammals, whereas ANP and ventricular NP (VNP) have been identified in eel hearts. Because VNP has also been found in the rainbow trout ventricle, we attempted to isolate NP from trout cardiac atria in order to determine whether ANP and VNP are common cardiac NPs in teleosts. In the present experiments, we isolated VNP and a novel atrial NP consisting of 29 amino acid residues from the atria. This new trout NP exhibited similar sequence identity to mammalian ANP and BNP (50–60%). Its homology to eel ANP was low (52%) compared with high homology of trout and eel VNP (78%). Based on yield, the content of this new NP in trout atria may be even smaller than that of VNP. The new trout atrial NP exhibited low relaxant activity in the chick rectum (only 1/10 of that of trout VNP), and extremely low vasorelaxant activity in the rat aortic strip (only 1/400 of that of human ANP). However, the new trout NP was equipotent with trout VNP and human ANP in relaxing trout epibranchial artery. Based on the sequence similarity with other NPs and on atrial content, the new NP isolated from trout atria cannot yet be assigned to a known member of the NP family.

Heavy and light chain variable region sequences and antibody properties of anti-phosphotyrosine antibodies reveal both common and distinct features.

Phosphotyrosine and similar analogs have been used to elicit antibodies that have found widespread use in the study of cellular tyrosine phosphorylation. In order to better understand the anti-phosphotyrosine immune response and to elucidate the details of the specific association between a tyrosine phosphate and an antibody combining site, we have undertaken a detailed comparison of antibody stability, specificity, apparent affinity, and primary structure for eight different anti-phosphotyrosine antibodies derived from immunizations with three different antigens. Two of these, 2G8 and 1G2, were derived from an immunization using azobenzylphosphonate conjugated to carrier, and five others, Py2, Py20, Py42, Py54, and Py69, were the products of an immunization with phosphotyrosine conjugated to carrier. Each of these anti-hapten antibodies was an IgG. One antibody, 129, an IgM, was the result of an immunization with a mixture of tyrosine-phosphorylated proteins which had been purified from growth factor treated cells. We found that antibody binding was significantly inhibited by millimolar levels of divalent cations or high concentrations of monovalent salt, with the exception of the antibody 129 where binding was significantly enhanced by both. Under optimal conditions, the highest apparent affinities for phosphotyrosine were observed for antibodies Py69 and Py20 (10(-6)-10(-7) M) and the lowest for 129 and 1G2 (10(-3)-10(-4). The heavy and light chain variable regions of seven of these antibodies were cloned and sequenced and a predominant anti-phosphotyrosine response was observed. The light chains of these antibodies could be assigned to one of two major VK groups, VK10 and VK19, with sequence identity between the different light chains of each class ranging from 65 to 100% at the amino acid level. Similar sequence identity was found among the heavy chain sequences (89-98% identity at the amino acid level) with the exception of one antibody, 2G8, which was only distantly related to the others (61-64% amino acid identity). These heavy chains belong to the same heavy chain family, J558. Two of the antibodies, Py20 and Py69, were clearly derived from the same progenitor cell since both share a highly unusual apparent V-D-D-JH organization. However, a significant level of somatic mutation has occurred between the two antibodies resulting in subtle changes in their apparent affinity and specificity.

Preferential association of kappa IIIb light chains with monoclonal human IgM kappa autoantibodies.

The predominance of the relatively uncommon V region subgroup isotype kappa III among the light chains of human monoclonal (IgM kappa) anti-IgG antibodies, (i.e., rheumatoid factors), was further documented through sequence analyses of ten such autoantibodies isolated from IgM-anti-IgG cold-insoluble immune complexes (mixed cryoglobulins). The amino-terminal sequence of all ten kappa-chains was characteristic for kappa III proteins and virtually identical to that of a prototype kappa III light chain. Similar sequence identity was found for kappa-chains isolated from three IgM kappa autoantibodies that formed cold-insoluble immune complexes with low-density lipoprotein (LDL). The thirteen light chains were found to be virtually identical in sequence for the first framework region (FR); ten of these proteins sequenced through the first complementarity-determining region (CDR) and into the second FR were markedly similar. The second CDR of five proteins was almost identical in sequence to that of the prototype kappa III-chain. Concordance was also demonstrated between the structural classification of the light chains as kappa III and their immunochemical classification as members of this V region subgroup. Serologic analyses of light chains isolated from seven IgM kappa autoantibodies (six anti-IgG, one anti-LDL) and of one intact IgM kappa anti-LDL antibody showed that each had antigenic determinants common to kappa II proteins. These light chains also expressed the antigenic determinant(s) of a V-region sub-subgroup of kappa III proteins designated kappa IIIb. Our studies confirm the preferential association of kappa III (and kappa IIIb) light chains with IgM kappa anti-IgG antibodies and demonstrate a similar association for IgM kappa anti-LDL antibodies. The finding that these and other types of IgM kappa autoantibodies, e.g., cold agglutinins, have remarkably similar light chains suggests an inherent restriction in the immune response to self-antigens.