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At least 109 records · Page 6Linked to original sources
PubMed2002

[Prospective analysis of conservative treatment of acute coronary syndrome].

The in-hospital and intermediate outcomes in 341 patients with acute coronary syndrome treated with conservative (non-invasive) interventions were analysed. The coronary reperfusion was achieved in 59% patients with acute myocardial infarction (MI) treated with thrombolysis, but in 27% patients with MI treated only heparin and aspirin. In-hospital survival was 91% in 93 patients with MI treated with thrombolysis and 82% in 71 patients with MI without thrombolysis. On 6 month survival was 86% in patients with MI treated with thrombolysis, 70% in patients with MI without thrombolysis, 79% in patients with non-Q-wave MI and 88% in patients with unstable angina.

Acute Disease↗
PubMed2001

Genetic analysis of conservation and variation of lipooligosaccharide expression in two L8-immunotype strains of Neisseria meningitidis.

Neisseria meningitidis strains A1 and M978 both express the lipooligosaccharide (LOS) L8 immunotype [Gu et al., J. Clin. Microbiol. 30 (1992) 2047-2053]. Under different growth conditions, strain A1 did not change its LOS profile whereas strain M978 produced variable LOS profiles on SDS-PAGE. To understand the genetic basis of LOS conservation and variation, their lgt locus encoding glycosyltransferases responsible for the biosynthesis of the alpha-chain of LOS was analyzed. Strain A1 possessed only two genes, lgtA and lgtH, at the lgt locus. The lgtA gene was inactivated due to a frameshift mutation; thus strain A1 expressed only L8 LOS. In contrast, strain M978 contained five genes lgtZ, lgtC, lgtA, lgtB and lgtE at this locus, thus it had a potential to express L1, L3,7 in addition to the L8 LOS. The data showed that strain A1 is a better reference strain for the L8 immunotype because of the stability of L8 LOS expression resulting from its unique lgt locus. In addition, these two strains had two new genetic organizations, lgtAH and lgtZCABE, compared to the reported gene organization at the lgt locus in N. meningitidis.

Bacterial Proteins↗
PubMed1994

Primer-terminus stabilization at the psi 29 DNA polymerase active site. Mutational analysis of conserved motif TX2GR.

The functional significance of the conserved motif TX2GR, included in one of the six main regions of amino acid sequence similarity identified in the C-terminal portion of both Escherichia coli DNA polymerase I-like and eukaryotic-type DNA polymerases (Blanco, L., Bernad, A., Blasco, M.A., and Salas, M. (1991) Gene (Amst.) 100, 27-38) has been studied by site-directed mutagenesis in the psi 29 DNA polymerase. A revised multiple alignment of this region, including 61 DNA polymerases belonging to these two superfamilies, is presented. In addition, based on amino acid sequence comparisons and by extrapolation to the crystal structure of T7 RNA polymerase, a similar motif (DX2GR) is predicted to be structurally and functionally equivalent in RNA polymerases, the other class of DNA-dependent polymerases. The severe defect in polymerization displayed by two of the psi 29 DNA polymerase mutants used in this study (T434N and R438I) is interpreted as the consequence of a decreased capacity to stabilize the binding of primer-template DNA structures in a polymerization-competent conformation. These mutants were also severely affected in the formation of terminal protein (TP)-dAMP initiation complex, a reaction in which psi 29 DNA polymerase is able to use the TP as primer.

Amino Acid Sequence↗
PubMed1997

Stabilization centers in proteins: identification, characterization and predictions.

Methods are presented to locate residues, stabilization center elements, which are expected to stabilize protein structures by preventing their decay with their cooperative long range interactions. Artificial neural network-based algorithms were developed to predict these residues from the primary structure of single proteins and from the amino acid sequences of homologous proteins. The prediction accuracy using only single sequence information is 65%, but the incorporation of evolutionary information in the form of multiple alignments and conservation scores raises the efficiency by 3%. The composition, relative accessibility, number and type of interactions, conservation and the X-ray thermal factor of the identified stabilization center residues are different, not only from the whole data set but from the rest of the long range interacting residues as well. The most frequent stabilization center residues are usually found at buried positions and have a hydrophobic or aromatic side-chain, but some polar or charged residues also play an important role in the stabilization. The stabilization centers show significant difference in the composition and in the type of linked secondary structural elements compared with the rest of the residues. The performed structural and sequential conservation analysis showed the higher conservation of stabilization centers over protein families. The relation of the proposed stabilization centers to folding nuclei is also discussed.

Algorithms↗
PubMed2001

Mutational analysis of conserved sequence motifs in the budding yeast Cdc6 protein.

The Cdc6 protein is required to load a complex of Mcm2-7 family members (the MCM complex) into prereplicative complexes at budding yeast origins of DNA replication. Cdc6p is a member of the AAA(+) superfamily of proteins, which includes the prokaryotic and eukaryotic clamp loading proteins. These proteins share a number of conserved regions of homology and a common three-dimensional architecture. Two of the conserved sequence motifs are the Walker A and B motifs that are involved in nucleotide metabolism and are essential for Cdc6p function in vivo. Here, we analyse mutants in the other conserved sequence motifs. Several of these mutants are temperature-sensitive for growth and are unable to recruit the MCM complex to chromatin at the restrictive temperature. In one such temperature-sensitive mutant, a highly conserved asparagine residue in the sensor I motif was changed to alanine. Overexpression of this mutant protein is lethal. This phenotype is very similar to the phenotype previously described for a mutation in the Walker B motif, suggesting a common role for sensor I and the Walker B motif in Cdc6 function.

Alleles↗
PubMed2006

Analysis of conserved glutamate residues in Porphyromonas gingivalis outer membrane receptor HmuR: toward a further understanding of heme uptake.

The aim of this study was to broaden the current knowledge about the Porphyromonas gingivalis heme receptor HmuR. Site-directed mutagenesis was employed to replace Glu427, Glu448, Glu458 and Glu503 by alanines and to construct a triple Glu427Ala/Glu448Ala/Glu 458Ala mutant. All iron/heme-starved P. gingivalis mutants showed decreased growth recovery when human serum as the iron/heme source was used, hmuR::ermF, hmuR (E503A) and hmuR (E427A,E448A,E458A) mutant strains being the most affected. E. coli cells expressing HmuR with mutated glutamate residues bound hemin, hemoglobin and hemin-serum albumin complex with the same efficiency as did the wild-type recombinant protein, suggesting that the residues were not directly involved in heme binding. These data indicate that in addition to two conserved histidine residues (His95 and His434), NPDL and YRAP motifs, conserved glutamate residues are important for HmuR to utilize heme present in serum hemoproteins.

Albumins↗
PubMed2002

Analysis of conserved and non-conserved amino acids critical for ALSV (Avian leukemia and sarcoma viruses) integrase functions in vitro.

Retroviral integrase (IN) is the viral enzyme responsible for the integration of viral DNA into host cellular DNA. In vitro, recombinant IN protein is able to catalyze the 3'-processing, strand transfer and disintegration activities. In order to analyze the importance of specific residues of ALSV (Avian leukemia and sarcoma viruses) IN protein, we introduced 31 amino acid substitutions either in residues previously shown by others to be involved in IN oligomerization or in selected conserved and non-conserved residues through the IN sequence. We tested, in vitro, the three catalytic activities of these mutants as well as their capacity to bind DNA. We found that (i) 88% of the substitutions occurring on well-conserved residues have an effect on IN activities (ii) two mutants (S85T in the central catalytic domain and N197C in the C-terminal domain) present a reduced efficiency of DNA binding compared to the wild type protein. Moreover, all mutations made on the dimer interface of C-terminal domain present reduced activities, suggesting an important role of this part of the protein. Finally, for some mutations, we observed differences between the ALSV and HIV (Human immunodeficiency virus) IN corresponding residues.

Alpharetrovirus↗
PubMed2000

The Internally Self-fertilizing Hermaphroditic Teleost Rivulus marmoratus (Cyprinodontiformes, Rivulidae) beta-Actin Gene: Amplification and Sequence Analysis with Conserved Primers.

To determine the ease and feasibility of amplifying the beta-actin gene in fish by the polymerase chain reaction (PCR), genomic DNAs of several fish (Rivulus, Southern top mouth minnow, common fat minnow, oily bitterling, carp, Far Eastern catfish, medaka, and European flounder) were extracted and used as a template with conserved primers, designed on the basis of high amino acid homology (approximately 98% or more). Among them, the self-fertilizing hermaphroditic fish Rivulus marmoratus was chosen for further characterization. After amplification of the Rivulus beta-actin PCR product with Taq polymerase, PCR product was subcloned to pCRII vector. After restriction enzyme mapping of Rivulus beta-actin gene, the amplified insert was sequenced using ALF Express automatic DNA sequencer with conserved internal primers. The R. marmoratus beta-actin gene consists of 1763 bp encoding 375 amino acids including 5 exons and 4 introns. The splicing and acceptance sites of the exon and intron boundaries of the Rivulus beta-actin gene were highly conserved with consensus sequences (GT/AG). The amino acid homology of R. marmoratus beta-actin to other species was high: 98.93% to human; 98.93%, Atlantic salmon; 98.93%, common carp; 98.93%, grass carp; 98.93%, zebrafish; 98.67%, medaka; and 98.40%, sea bream. To determine the expression of the R. marmoratus beta-actin gene in liver and ovary, reverse transcriptase-polymerase chain reaction was carried out with internal primers. In conclusion, these universal primers are successful in the rapid cloning of the fish beta-actin gene by PCR, based on a high homology of the beta-actin gene conserved through evolution. This approach will be applicable to the isolation of other beta-actin homologues in the investigation of phylogenetic comparisons of fish species, along with a possible application to cloning strategy in other conserved genes.

Journal Article↗
PubMed2005

Genetic analysis and conservation of the endangered Canary Island woody sow-thistle, Sonchus gandogeri (Asteraceae).

Sonchus gandogeri, a woody sow-thistle, is an endangered Canary Island endemic with only two known populations, one in the El Golfo and another in the Las Esperillas of El Hierro. Amplified fragment length polymorphism (AFLP) markers were used to assess the genetic variation within and among populations. The mean genetic diversity of two populations was estimated to be 0.380, and the El Golfo population (0.380) had higher genetic diversity than the southeastern one (0.268). The unbiased Nei's genetic identity between the two populations was 0.846. The mean genetic diversity of S. gandogeri was much higher than that of the other endangered plant species. This is perhaps due to breeding system, life form, extinction, and/or introgressive hybridization and hybrid origin of the taxon. This study also indicates that the two populations are not strongly differentiated (G(ST)=0.149). This study suggests that S. gandogeri is more likely to become extinct due to environmental or demographic forces than genetic factors, such as inbreeding depression. More strict control of introduced herbivores is necessary to protect these populations, and germplasm collection for ex situ conservation is needed.

Atlantic Islands↗
PubMed2026

Comparative analysis of conserved non-coding elements identifies gene regulatory networks rewired during the water-to-land transition in vertebrates.

The conquest of land by vertebrates has been a pivotal moment in evolutionary history. Adapting to the new habitats necessitated numerous changes in vertebrate anatomy and physiology, creating an enduring imprint on the developmental gene regulatory networks (GRNs) of tetrapods. The increase of high-quality genomic resources over the past decade has made it possible to study the genomic legacy of the water-to-land transition. While much attention has been given to the highly conserved non-coding elements (CNEs) of the genome that share high levels of similarity across evolutionarily diverged clades, recent evidence suggests that perhaps comparable attention should be given to "missing" CNE-s, conserved sequence patches present in extant stem gnathostomes and actinopterygian fishes that have become undetectable in tetrapods during the adaptation to terrestrial life, whether through true sequence loss or divergence beyond alignability. These sequences could help us reveal the relaxation of certain developmental constraints, related to the aquatic lifestyle, that made reaching new adaptive peaks in the developmental landscape possible. In this paper, we search for such CNEs and characterize them in comparison with pan-Gnathostome CNEs, using the zebrafish (Danio rerio) genome as a reference. Our results suggest that the rewiring of developmental networks related to pigmentation and muscle structure formation has left the largest genomic imprint. We also find that components of canonical Wnt and Hedgehog signalling, are enriched among CNEs retained in fish.

cis-regulatory evolution↗
PubMed1994

Functional analysis of conserved cysteine residues in the catalytic subunit of the yeast vacuolar H(+)-ATPase.

The A subunit of the yeast vacuolar ATPase contains three highly conserved cysteines: Cys-261, Cys-284, and Cys-538. Cys-261 is located within the nucleotide-binding P-loop. Each of the conserved cysteines, and one nonconserved cysteine, Cys-254, were altered to serine by site-directed mutagenesis, and the effects on growth at pH 7.5 were determined. The Cys-254-->Ser, Cys-261-->Ser and the double mutants all grew at pH 7.5 and contained nitrate- and bafilomycin-sensitive ATPase activity. However, the ATPase activities of the Cys-261-->Ser and the double mutants were insensitive to the sulfhydryl group inhibitor, N-ethylmaleimide, demonstrating that Cys-261 is the site of inhibition by N-ethylmaleimide. Changing either Cys-284 or Cys-538 to serine prevented growth at pH 7.5. Cys-284 and Cys-538 thus appear to be essential cysteine residues which are required either for assembly or catalysis.

Base Sequence↗
PubMed1987

Molecular characterization of human immunodeficiency virus from Zaire: nucleotide sequence analysis identifies conserved and variable domains in the envelope gene.

To examine the genetic relatedness of human immunodeficiency viruses (HIV) from different geographic locations, we molecularly cloned the genome of HIV isolated from a Zairian AIDS patient. Restriction mapping of the recombinant clone, designated HIV-Zr6, revealed both common (as observed in other HIV isolates) and unique restriction sites. The DNA clone of HIV-Zr6, shown to give rise to infectious cytopathic virus after transfection of cultured lymphoid cells, was sequenced in several regions. The long terminal repeat (LTR), open reading frame 1 (ORF1), C-terminal envelope (env) gene domain, and ORF2 showed less than 6% difference in nucleotide sequence when compared to other HIV isolates including human T-lymphotropic virus-type III (HTLV-III) clone B10, lymphadenopathy-associated virus-1 (LAV-1), and AIDS-associated retrovirus-2 (ARV-2). About 15% difference in nucleotide sequences was noted in the N-terminal env gene domain. Alignments of env gene sequences revealed conserved, moderately variable, and hypervariable stretches in the predicted amino acid sequences. This model provides a basis for assessing the significance of sequence variation on properties controlled by the viral Env glycoproteins such as cell tropism and immunogenicity.

Acquired Immunodeficiency Syndrome↗
PubMed1994

The thiol-specific antioxidant protein from human brain: gene cloning and analysis of conserved cysteine regions.

The complete cDNA encoding human thiol-specific antioxidant protein (PRP) was isolated from a human brain cDNA library in the lambda Zap expression vector. An open reading frame (ORF) was identified and found to encode a polypeptide of 197 aa with a M(r) of 21,729. The cDNA contained 98 bp of 5'-untranslated sequence (UTR) and 259 bp of 3'-UTR containing a poly(A) signal, AATAAA. Expression of the human PRP cDNA in Escherichia coli yielded a functionally active protein. The observed local sequence homologies between human PRP and other homologous proteins whose functions have not yet been defined give important insight into elucidating the biochemical function of a new protein family which has highly conserved regions containing cysteine.

Amino Acid Sequence↗
PubMed2004

Mutational analysis of conserved AAA+ residues in the archaeal Lon protease from Thermoplasma acidophilum.

The Lon protease from the archaeon Thermoplasma acidophilum (TaLon) is composed of an N-terminal ATPase associated with various cellular activities (AAA+) domain and a C-terminal Lon protease domain. Although related in sequence to the soluble Lon proteases, TaLon was shown to be membrane-bound in its native host and also when expressed in Escherichia coli. Recombinant TaLon was purified as a functional high-molecular weight complex displaying ATPase and proteolytic activity. Mutagenesis of conserved AAA+ residues revealed that the Walker A and B motifs, and the sensor 1 and sensor 2' residues were essential for the ATPase activity, while the sensor 2 and the arginine finger were involved in activation of the protease domain.

ATP-Dependent Proteases↗
PubMed2003

Analysis of conserved residues of the human puromycin-sensitive aminopeptidase.

The puromycin-sensitive aminopeptidase (ApPS) is a zinc metallopeptidase involved in the degradation of neuropeptides. Putative catalytic residues of the enzyme, Cys146, Glu338, and Lys396 were mutated, and the resultant mutant enzymes characterized. ApPS C146S exhibited normal catalytic activity, ApPS E338A exhibited decreased substrate binding, and ApPS K396I exhibited decreases in both substrate binding and catalysis. ApPS K396I and ApPS Y394F were analyzed with respect to transition state inhibitor binding. No effect was seen with the K396I mutation, but ApPS Y394F exhibited a 3.3-fold lower affinity for RB-3014, a transition state inhibitor, indicating that Tyr394 is involved in transition state stabilization.

Amino Acid Sequence↗
PubMed2003

Quality control of 2721 cornea transplantations of issued by Center of Corneal Analysis and conservation by the Bank of Tissue and Cells of Tuscany.

Knowledge of the clinical evolution of corneal transplantation is fundamental to its improvement by upgrading the modalities and conservation methods in eye banking. The purpose of this retrospective study was to review of the results obtained with corneal transplantation performed at 50 Italian hospitals supplied by our center. From July 1997 through December 2002, 2721 corneas were used for transplantation. A questionnaire was mailed to every transplant center, that included queries about the number of corneal transplants performed, the number of primary and late failures, and the prevalence of posttransplant endopthalmitis. Preliminary results revealed no correlation between primary/late failures and the morphologic properties of the cornea, donor age, although a few failures seemed related to the pathology of the recipient.

Aged↗