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R L Rodriguez

Publications and source records attributed to R L Rodriguez.

At least 37 records · Page 2Linked to original sources
PubMed1990

The isolation and characterization of a barley 1,3-1,4-beta-glucanase gene.

The barley gene encoding isozyme I of 1,3-1,4-beta-glucanase was isolated and sequenced. The 6260-bp region sequenced included 1885 bp of the 5'-flanking region, the entire coding region, an intron of 2490 bp, and 792 bp of the 3'-flanking region. The 1,3-1,4-beta-glucanase mRNA was found to be regulated at the level of RNA accumulation by both gibberellins (positively) and abscisic acid (negatively) in barley aleurones. The mRNA for isozyme II preferentially accumulated (70%) relative to the mRNA for isozyme I (30%) in poly(A)-rich RNA isolated from material including both the aleurone and the scutellum tissues. The gene family encoding 1,3-1,4-beta-glucanase enzymes in barley was found to be comprised of two closely related genes, isozymes I and II, as well as several related sequences that could be identified by Southern blot analysis. The nucleotide sequence for the 5' untranslated leader and the coding region for the signal peptide of the isozyme II transcript were determined from a cDNA produced by the polymerase chain reaction. The structure of the protein encoded by the isozyme I gene is also discussed.

Abscisic Acid↗
PubMed1990

Structural organization and differential expression of rice alpha-amylase genes.

Rice alpha-amylases are encoded by a multigene family that has previously been classified into 5 hybridization groups. DNA sequence and Southern blot analysis identified three genes (RAmy1A, RAmy1B and RAmy1C) in Group 1 with DNA sequence identity of at least 90%. Hybridization Group 2 is represented by only one gene, RAmy3D, which is identical to a previously characterized cDNA, pOS137. RAmy3D is physically linked to the sole representative of Group 5, RAmy3E. The identity between these two genes is 81.4% in the coding region but less than 50% in the 5' and 3' flanking regions. Northern blot analysis and RNA-PCR were used to detect the expression of alpha-amylase genes in various tissues. Alpha-amylase mRNA was abundant in germinating seeds and callus. Some genes were also expressed at much lower levels in roots, young leaves and immature seeds. RAmy1A and RAmy3E were expressed in all tissues while RAmy3D was expressed in all tissues except the immature seeds. RAmy1B was weakly expressed only in callus. RAmy1A transcript was most abundant in the germinating seeds, while RAmy3D and RAmy3E transcripts were most abundant in callus and immature seeds, respectively.

Amino Acid Sequence↗
PubMed1990

Expression and secretion of rice alpha-amylase by Saccharomyces cerevisiae.

We report the high level expression and secretion of rice alpha-amylase isozyme by Saccharomyces cerevisiae. Transcription of this gene was under control of the yeast enolase promoter. The synthesized protein had an approximate molecular size of 45 kDa and a pI of approx 4.7 to 5.0. The rice alpha-amylase signal peptide was recognized and efficiently processed by yeast and the active, glycosylated enzyme was secreted into the culture media. This enzyme was purified to homogeneity by affinity chromatography and its enzymatic properties were characterized. The Km and Vmax were found to be similar to those of alpha-amylases from other organisms. The high level of secretion observed in these studies may be due to the unique features of the rice signal peptide and/or to the glycosylation of the recombinant enzyme.

Amino Acid Sequence↗
PubMed1990

Classification and characterization of the rice alpha-amylase multigene family.

To establish the size and organization of the rice alpha-amylase multigene family, we have isolated 30 alpha-amylase clones from three independent genomic libraries. Partial characterization of these clones indicates that they fall into 5 hybridization groups containing a total of 10 genes. Two clones belonging to the Group 3 hybridization class have more than one gene per cloned fragment. The nucleotide sequence of one clone from Group 1, lambda OSg2, was determined and compared to other known cereal alpha-amylase sequences revealing that lambda OSg2 is the genomic analog of the rice cDNA clone, pOS103. The rice alpha-amylase genes in Group 1 are analogous to the alpha-Amy1 genes in barley and wheat. lambda OSg2 contains sequence motifs common to most actively transcribed genes in plants. Two consensus sequences, TAACAAGA and TATCCAT, were found in the 5' flanking regions of alpha-amylase genes of rice, barley and wheat. The former sequence may be specific to alpha-amylase gene while the latter sequence may be related to a 'CATC' box found in many plant genes. Another sequence called the pyrimidine box (TCCTTTTTC) was found in the alpha-amylase genes as well as other genes regulated by gibberellic acid (GA). Comparisons based on amino acid sequence alignment revealed that the multigene families in rice, barley and wheat shared a common ancestor which contained three introns. Some of the descendants of the progenitor alpha-amylase gene appear to have lost the middle intron while others maintain all three introns.

Amino Acid Sequence↗
PubMed1990

The alpha-amylase genes in Oryza sativa: characterization of cDNA clones and mRNA expression during seed germination.

Two cDNA clones, pOS103 and pOS137, were isolated which code for distinct alpha-amylase isozymes in germinating rice seeds. Sequence analysis indicated that the clones encode polypeptides of approximately 48 kDa, both of which possess a signal peptide involved in directing secretion of the protein. Comparison of the two rice alpha-amylase amino acid sequence showed that they are 76% similar to each other, while showing 85% to 90% similarity with other cereal alpha-amylases. A comparison of eleven cereal alpha-amylases also revealed three new conserved regions (I', II', and IV') not previously identified in the animal, bacterial, and fungal alpha-amylases. Regions I' and IV' are sites for intron splicing while region II' is probably involved in calcium binding. One of the rice alpha-amylase cDNAs, pOS103, encodes a protein that has two potential N-glycosylation sites, one in the signal peptide and the other in the mature portion of the protein. The cDNA clone, pOS137, encodes an alpha-amylase with a single glycosylation site in the signal peptide, suggesting that the mature OS137 isozyme is not glycosylated. Analysis of the expression of these genes in germinating rice seeds indicated that mRNA corresponding to pOS103 and pOS137 could be detected throughout a 48 h period of seed imbibition. RNA levels, however, were dramatically stimulated by treatment of embryoless half-seeds with exogenous GA3. Our results demonstrate that at least two forms of alpha-amylase are expressed in germinating rice seeds and that the expression of these genes is regulated by the phytohormone GA3.

Amino Acid Sequence↗
PubMed1986

Selective management of arterial occlusion with low-dose streptokinase.

Fibrinolytic therapy has been reported as a limited aid in limb salvage. It is allegedly fraught with complications and pitfalls; however, the usage of low-dose streptokinase has never totally been explored. The purpose of this study was to examine the role of low-dose streptokinase in arterial occlusion. Nineteen patients (12 men and 7 women) with an average age of 68 years (range 37 to 79 years) and a total of 22 limbs under study were treated with low-dose streptokinase (initial intravenous dose of 10,000 units and a subsequent dose of 5,000 to 10,000 units/hour) for 1 to 3 days. Seven of the patients underwent further operative intervention and four underwent angioplasty. Seven patients had diabetes controlled by insulin. Results were assessed clinically by angiography and by noninvasive measurement of distal pressure. Within 30 days of low-dose streptokinase therapy, four patients died from a myocardial infarction that was thought to be part of their total disease process. Limited salvage occurred in 19 of 22 limbs at risk. Ten patients (12 extremities) underwent subsequent balloon angioplasty or bypass surgery. There were three amputations. One patient received a below-knee amputation rather than an expected above-knee amputation, one had a below-knee amputation, and one had an above-knee amputation. In all cases, the patency or lack thereof was confirmed by angiography. The mean ankle to brachial pressure indices increased from 0.07 +/- 0.15 to 0.64 +/- 0.14 after therapy. There were no prolonged hospital stays due to complications in any of the patients. Selective infusions of low-dose streptokinase lyse thrombi, open collaterals, and provide an opportunity for extended surgery and angioplasty, increasing limb salvage without the risks of coagulopathy and bleeding that may occur in full-dose thrombolysis.

Adult↗
PubMed1985

The aminoglycoside-resistance operon of the plasmid pSa: nucleotide sequence of the streptomycin-spectinomycin resistance gene.

The nucleotide sequence of the probable C terminus of the kanamycin-resistance gene (KmR) and the probable complete sequence of the streptomycin-spectinomycin-resistance gene (SpR) of the IncW plasmid pSa have been determined. The two genes appear to be oriented in the same direction and separated by a spacer region of 53 bp, with transcription proceeding from the KmR gene into the SpR gene. An RNA transcript encompassing the C terminus of the KmR gene, the 53-base spacer, and the N terminus of the SpR gene has the potential to form a stem-loop structure with a free energy value of -68 kcal/mol. The SpR gene of pSa has extensive sequence homology with the aadA gene of the plasmid R538-1. Comparison of the proposed amino acid sequence of the KmR protein of pSa with those of two aminoglycoside phosphotransferases revealed a region of potential homology with those proteins.

Base Sequence↗
PubMed1985

Mutations that affect the translation efficiency of Tn9-derived cat gene in Bacillus subtilis.

We have isolated two spontaneous mutations that increase the expression of the Tn9-derived cat gene in Bacillus subtilis. These mutations, which appear to affect initiation of translation of chloramphenicol acetyltransferase (CAT; acetyl-CoA:chloramphenicol 3-O-acetyltransferase, EC 2.3.1.28) consist of a tandem duplication and triplication of a 55-base-pair sequence located at the 5' end of cat. Included in the repeated sequence are the Shine-Dalgarno site, initiation codon, and a region of dyad symmetry located within the structural portion of the cat gene. A striking feature of the mutated initiation sites is their potential to form stem-loop structures at the 5' end of the cat messenger RNA. Within the single-stranded loops of these structures are the ribosome binding site and initiation codon for the cat gene. It appears that the Gram-negative cat translation initiation site has mutated to permit efficient utilization in B. subtilis without directly affecting Shine-Dalgarno sequence homology. This report suggests that secondary structure in the vicinity of the Shine-Dalgarno site can exert a strong positive influence on the initiation of translation in B. subtilis.

Acetyltransferases↗
PubMed1984

Benzyl derivative facilitation of transcription in Escherichia coli at the ara and lac operon promoters: metabolite gene regulation (MGR).

A number of benzyl derivatives have been tested for their ability to induce the expression of the araBAD operon in an Escherichia coli K-12 strain. Those derivatives shown to be stimulatory include: benzoic acid (BA), para-amino benzoic acid (PABA), para-hydroxy benzoic acid (PHBA), ortho-amino benzoic acid (OABA), 3-hydroxy-4-methoxy phenylethylamine (MTA), and 4-hydroxy-3-methoxyphenol acetic acid (HVA). The araC gene product was necessary to facilitate the induction. To further characterize if the inductive effect was mediated at the level of transcription, an araBAD-tetracycline resistant (Tcr) operon fusion plasmid (pAP-B) was employed. Benzyl derivatives which induce expression of the araBAD operon in situ also induced a Tcr phenotype with pAP-B. Both indole acetic acid (IAA) and imidazole (IM), which were previously shown to circumvent the necessity for cAMP in the induction of the araBAD operon, also induced a Tcr phenotype with pAP-B. Induction of lac or other cAMP responding operons with the inducing molecules at the chromosomal level was not detectable when assessed by carbon utilization. However, a lacZYA-Tcr operon fusion plasmid (pLPI) did respond to IAA and several of the inducing benzyl derivatives. Catabolite repression of chromosomal araBAD expression was reversed when the exogenous concentration of OABA was elevated. Similar effects on the Tcr phenotypes conferred by pAP-B and pLP1 were observed when OABA or several other inducing benzyl derivatives were present exogenously.

Aldose-Ketose Isomerases↗
PubMed1984

Purification of restriction endonuclease XcyI from Xanthomonas cyanopsidis.

A new Type II restriction endonuclease XcyI, purified from Xanthomonas cyanopsidis 13D5, is an isoschizomer of SmaI and XmaI that cleaves at the nucleotide sequence 5'-C decreases CCGGG-3' of double-stranded DNA. The single restriction activity present in this strain permits rapid purification of 8000 units of cleavage activity from 10 g of freshly harvested cells. The resulting XcyI preparation is free of contaminating nuclease activities that interfere with in vitro manipulation of DNA.

Bacterial Proteins↗
PubMed1984

Nucleotide sequence of the 3' terminal region of the LEU2 gene from Saccharomyces cerevisiae.

The 3' terminal region from the LEU2 gene of yeast has been sequenced. Two open reading frames (ORF) have been identified, one of which constitutes the 3' terminus of beta-isopropylmalate dehydrogenase, the product of the LEU2 gene. A noncoding spacer region located between the ORFs contains two consensus-type transcriptional terminators. The terminator-like sequences are oriented in opposing directions on opposite DNA strands. The noncoding spacer region may represent a single terminator for the LEU2 gene or two separate terminators involved in blocking convergent transcription from an as yet unidentified yeast gene.

3-Isopropylmalate Dehydrogenase↗
PubMed1984

Histidine operon control region of Klebsiella pneumoniae: analysis with an Escherichia coli promoter-probe plasmid vector.

The control region for the histidine operon of Klebsiella pneumoniae was cloned and analyzed with the Escherichia coli promoter-probe plasmid pPV33. A restriction fragment which contained the his control region was identified by its ability to activate the tetracycline resistance (Tcr) gene on this vector. Expression of Tcr by bacteria containing the his promoter-active plasmid was found to be under the attenuation control of the his promoter. DNA sequence analysis of the his control region revealed a base sequence homology of approximately 86% of the analogous DNA sequences of E. coli and Salmonella typhimurium. Most of the base alterations in the K. pneumoniae DNA sequence were found to reside in regions flanking the transcriptional and translational regulatory sites.

Alcohol Oxidoreductases↗
PubMed1983

A comparison of the origin of replication of pSa with R6K.

The plasmid pOri3 is a derivative of the origin of replication of pSa. Replication is defective as a result of a truncated repA gene, the product of which is required for plasmid replication. The defective replication is complemented by the presence of the intact repA gene of pSa, or by the presence of the plasmid R6K. The basis of this complementation has been examined by comparing the nucleotide sequence of the origin of pSa with that of R6K. A 13 base pair sequence present twice in the origin of pSa has homology with a 13 base pair sequence that is present fourteen times in the origin of R6K. These sequences may be the binding sites for the initiator proteins of these two plasmids. The location of these binding sites relative to the genes for the initiator proteins suggests that an autoregulatory control mechanism for the synthesis of the initiator proteins may also play a role in the control of plasmid copy number.

Base Sequence↗
PubMed1983

M13 bacteriophage and pUC plasmids containing DNA inserts but still capable of beta-galactosidase alpha-complementation.

A DNA fragment encoding the transposon Tn9 chloramphenicol acetyltransferase gene (cat) was inserted into M13 phage and pUC plasmid cloning vehicles. When the cat gene was inserted in the same orientation as the lacZ gene, two new polypeptides were produced. One polypeptide possessed chloramphenicol acetyltransferase activity, while the other expressed beta-galactosidase alpha-donor activity. Both new polypeptides were translated from a hybrid messenger RNA initiating from the lac promoter. These observations may help explain why not all inserts produce white plaques.

Acetyltransferases↗
PubMed1983

Nucleotide sequence of the promoter and amino-terminal coding region of the glutamate dehydrogenase structural gene of Escherichia coli.

A 610-bp DNA fragment carrying the promoter and amino-terminal coding regions of the glutamate dehydrogenase (GDH) structural gene from Escherichia coli has been sequenced. The amino-terminal sequence of the enzyme was also determined to help localize the transcriptional and translational signals for this gene. Three possible promoters and a CRP binding site were identified by concensus criteria. The sequence of 102 amino acids at the amino terminus of the enzyme is compared with the amino acid sequence from other GDH enzymes.

Animals↗