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C Lemieux

Publications and source records attributed to C Lemieux.

At least 73 records · Page 4Linked to original sources
PubMed1991

Cloning and characterization of the Chlamydomonas moewusii mitochondrial genome.

We report that the mitochondrial genome of Chlamydomonas moewusii has a 22 kb circular map and thus contrasts with the mitochondrial genome of Chlamydomonas reinhardtii, which is linear and about 6 kb shorter. Overlapping restriction fragments spanning over 90% of the C. moewusii mitochondrial DNA (mtDNA) were identified in a clone bank constructed using a Sau3AI partial digest of a C. moewusii DNA fraction enriched for mtDNA by preparative CsCl density gradient centrifugation. Overlapping Sau3AI clones were identified by a chromosome walk initiated with a clone of C. moewusii mtDNA. The mtDNA map was completed by Southern blot analysis of the C. moewusii mtDNA fraction using isolated mtDNA clones. Regions that hybridized to C. reinhardtii or wheat mitochondrial gene probes for subunit I of cytochrome oxidase (cox1), apocytochrome b (cob), three subunits of NADH dehydrogenase (nad1, nad2 and nad5) and the small and the large ribosomal RNAs (rrnS and rrnL, respectively) were localized on the C. moewusii mtDNA map by Southern blot analysis. The results show that the order of genes in the mitochondrial genome of C. moewusii is completely rearranged relative to that of C. reinhardtii.

Animals↗
PubMed1991

A group I intron in the chloroplast large subunit rRNA gene of Chlamydomonas eugametos encodes a double-strand endonuclease that cleaves the homing site of this intron.

During interspecific crosses between Chlamydomonas eugametos and Chlamydomonas moewusii, an optional group I intron of 955 base pairs (CeLSU.5) in the C. eugametos chloroplast large subunit rRNA gene undergoes a duplicative transposition event which is associated with frequent co-conversion of flanking cpDNA sequences. In the present study, we show that the basic protein of 218 amino acids encoded by CeLSU.5 could mediate the phenomenon of intron transposition, also called intron homing. We overexpressed the ORF specifying this protein in E. coli using expression vectors that contain a C. moewusii cpDNA sequence encompassing the intron homing site. The expression product was found to exhibit a double-strand DNA endonuclease activity that is specific for the homing site. This activity was detected in vivo by self-linearization of the expression plasmids.

Amino Acid Sequence↗
PubMed1991

Conformational restriction of the phenylalanine residue in a cyclic opioid peptide analogue: effects on receptor selectivity and stereospecificity.

In an effort to determine the effect of side chain conformational restriction on opioid receptor selectivity, the cyclic phenylalanine analogues 2-aminoindan-2-carboxylic acid (Aic), 2-aminotetralin-2-carboxylic acid (Atc), and tetrahydroisoquinoline-3-carboxylic acid (Tic) were substituted for Phe in the potent cyclic opioid peptide analogue H-Tyr-D-Orn-Phe-Glu-NH2, which lacks significant opioid receptor selectivity. Compounds were tested in mu- and delta-opioid receptor representative binding assays and bioassays in vitro. The analogue H-Tyr-D-Orn-Aic-Glu-NH2 was found to be a potent agonist with high preference of mu receptors over delta receptors. Opening of the five-membered ring of Aic in the latter peptide, as achieved through substitution of C alpha-methylphenylalanine or o-methylphenylalanine, resulted in only slightly selective compounds, indicating that the high mu selectivity of the Aic analogue is exclusively the consequence of the imposed side chain conformational restriction. Both diastereoisomers of H-Tyr-D-Orn-(D,L)-Atc-Glu-NH2 were highly mu-selective and, in contrast to the weak affinity observed with the D-Phe3 analogue as compared to the L-Phe3 analogue, both had similar potency. Thus, stereospecificity was lost as a consequence of side chain conformational restriction. Further structure-activity data obtained with analogues containing L- or D-homophenylalanine (Hfe) or 3-(1'-naphthyl)alanine (Nap) in place of Phe3 and consideration of geometric interrelationships between Nap and the L and D isomers of Atc, Hfe, and Phe led to the proposal that the D-Phe3 and the D-Atc3 analogue may have different modes of binding to the receptor. The very low potency observed with H-Tyr-D-Orn-N alpha MePhe-Glu-NH2 (N alpha MePhe = N alpha-methylphenylalanine) and H-Tyr-D-Orn-Tic-Glu-NH2 indicated that N alpha-alkylation at the 3-position is detrimental to activity.

Amino Acid Sequence↗
PubMed1991

Direct electrophilic fluorination of tyrosine in dermorphin analogues and its effect on biological activity, receptor affinity and selectivity.

In a preliminary communication we reported [(Tetrahedron Lett. 31, 619 (1990)] that acetyl hypofluorite can be used efficiently to introduce fluorine regiospecifically (ortho to OH) into the phenolic ring of tyrosine-containing peptides. This procedure has been applied to the fluorination of a number of mu-selective opioid peptides derived from dermorphin. While the procedure can be used even when the side chains of Arg, Lys, and Tyr are left unprotected, the sulfoxide of a Met(O)-containing analogue was oxidized to sulfone faster than fluorination of the phenolic ring. This method can also be used when the peptide is attached to Merrifield resin. Thus, Tyr(3-F)-D-Ala-Phe-Gly-NH2 and Tyr(3-F)-D-Arg-Phe-Lys-NH2 (F-DALDA) have been prepared, purified, and characterized. Affinities of these fluorinated peptides for both mu- and delta- opioid receptors are reduced (two- to nine-fold) relative to their nonfluorinated analogues, but their selectivity for mu-opioid receptors is not significantly altered. Similarly, the in vitro biological potencies (GPI and MVD assays) of the fluorinated analogues are reduced (two- to seven-fold) relative to their nonfluorinated parent peptides. Thus, F-DALDA, which has high affinity (Ki mu = 15.2 nM) and selectivity (Ki delta/Ki mu = 5390) for mu-opioid receptors, has potential use in biochemical studies which utilize 19F or 18F- labeled compounds.

Amino Acid Sequence↗
PubMed1990

Collaborative evaluation of antigen detection by a commercial latex agglutination test and enzyme immunoassay in the diagnosis of invasive candidiasis.

The Cand-Tec Candida detection system and enzyme immunoassay for serum mannan were retrospectively compared in a controlled collaborative evaluation of antigen detection in 32 patients with candidiasis proven by biopsy or culture from a normally sterile site and with sera drawn within 7 days of inclusion. With a threshold titer of 1/8, which excluded false-positive results in 17 hospitalized patients without candidiasis, sensitivities for all 32 patients with candidiasis were 44% for the Cand-Tec assay and 17% for the enzyme immunoassay. Both assays provided greater sensitivity when sera were drawn within 24 h of inclusion in the study and in the category of patients with invasive candidiasis (57% by Cand-Tec and 33% by enzyme immunoassay). The Cand-Tec assay gave false-positive results (titer, greater than or equal to 1/8) in 4 of 6 patients with transient candidemia, in 1 of 20 otherwise healthy patients with rheumatoid factor, and in 1 patient with a positive cryptococcal latex agglutination test. Three serum specimens from 3 of 32 patients with candidiasis contained rheumatoid factor and gave titers of greater than or equal to 1/8 by the Cand-Tec assay. Detection of serum mannan by enzyme immunoassay was less sensitive but more specific than the Cand-Tec Candida detection system for the diagnosis of invasive candidiasis.

Antigens, Fungal↗
PubMed1990

Metabolic characteristics of cat kidney: failure to adapt to metabolic acidosis.

During studies performed on domestic cats made acidotic with ammonium chloride, it was found that the cat kidney is unable to adapt to metabolic acidosis. Renal proximal tubules do not increase their production of ammonia or glucose from glutamine during acidosis. During in vivo studies, the renal excretion of ammonia did not change much during acidosis. Other metabolic parameters in the cat were not very different from those found in other animals such as rat or dog. However, it was found that cats may show a relatively high plasma glucose concentration compared with other animals. Plasma insulin concentration was normal, and the animals showed no evidence of diabetes mellitus. It is not known whether limitation of ammoniagenesis and elevated plasma glucose concentration also characterize larger felidae such as panthers and cougars.

Acid-Base Equilibrium↗
PubMed1989

Mobile introns: definition of terms and recommended nomenclature.

A number of introns in mitochondrial, chloroplast, nuclear or prokaryotic genes have recently been shown to encode double-strand sequence-specific endonucleases. Such introns are mobile genetic elements that insert themselves at or near the cleaved sites. A uniform nomenclature to designate the molecular elements involved in the phenomenon of intron mobility is proposed.

Base Sequence↗
PubMed1989

Two group I introns with long internal open reading frames in the chloroplast psbA gene of Chlamydomonas moewusii.

We report the nucleotide sequence of the chloroplast psbA gene encoding the 32 kilodalton protein of photosystem II from Chlamydomonas moewusii. Like its land plant homologues, this green algal protein consists of 353 amino acids. The C. moewusii psbA gene is composed of three exons containing 252, 11 and 90 codons and of two group I introns containing 2363 and 1807 nucleotides. Each of the introns features an internal open reading frame (ORF) that potentially encodes a basic protein of more than 300 residues. The primary sequences of the putative intron-encoded proteins are unrelated and none of them shares conserved elements with any of the proteins predicted from the group I intron sequences published so far. The first C. moewusii intron is inserted at the same position as the fourth intron of the psbA gene from Chlamydomonas reinhardtii; the second intron lies at a novel site downstream of this position. On the basis of their RNA secondary structures, the C. moewusii introns 1 and 2 can be assigned to subgroups IA and IB, respectively. However, intron 1 is not typical of subgroup IA introns, its most unusual feature being the location of the ORF in the "loop L5" region. To our knowledge, this is the first time that an ORF is located in this region of the group I intron structure.

Amino Acid Sequence↗
PubMed1989

An optional group I intron between the chloroplast small subunit rRNA genes of Chlamydomonas moewusii and C. eugametos.

We report the presence of a 402 bp group I intron in the chloroplast small subunit (SSU) rRNA gene of Chlamydomonas moewusii. The intron in inserted within the highly conserved '530 loop', at a site corresponding to positions 531-532 of the E. coli 16rRNA. Residues surrounding the insertion site almost certainly play an important role in ribosomal proofreading function as they proved to be protected by tRNAs in E. coli 16S rRNA (Moazed and Noller 1986; Stern et al. 1986). The C. moewusii intron revealed a secondary structure model which differs substantially from those of the typical subgroup IA and IB introns. This model, however, shows striking similarities with the structures of the C. reinhardtii chloroplast 23S rRNA gene intron (Rochaix et al. 1985), the S. cerevisiae mitochondrial COB3 intron (Holl et al. 1985) and the three introns of phage T4 in the nrdB, td and sunY genes (Shub et al. 1988). The SSU rRNA gene intron is absent from C. eugametos, an alga that is interfertile with C. moewusii. The presence/absence of the intron account for a 390 bp restriction fragment length polymorphism between the two algal SSU rRNA genes, a polymorphic locus that is strictly co-inherited with a tightly linked streptomycin resistance mutation (sr-2) in interspecific hybrids between the two algae.

Base Sequence↗
PubMed1989

Cyclic enkephalin analogues containing alpha-amino-beta-mercapto-beta,beta-pentamethylenepropionic acid at positions 2 or 5.

Analogues of the highly potent and delta-receptor-selective enkephalins 1-4 were prepared with alpha-amino-beta-mercapto-beta,beta-pentamethylenepropionic acid (Apmp) replacing the beta,beta-dimethylcysteine (Pen) at positions 2 or 5. The peptides 5-8 were prepared by employing D,L-Apmp and, following oxidative cyclization, the resulting diastereomeric peptides were separated and purified by preparative high performance liquid chromatography. Compounds 7 and 8, with D- or L-Apmp substituted at position 5 are approximately 5 orders of magnitude more potent in the MVD assay than analogues 5 or 6 with D- or L-Apmp at position 2. While displaying less delta-receptor selectivity than the corresponding Pen-containing compounds, 7 and 8 are an order of magnitude more potent. All the analogues showed diminished delta-receptor selectivity in the rat brain binding assay. Compounds 7 and 8 displayed delta-receptor affinity comparable to the corresponding Pen-containing analogues.

Amino Acids, Sulfur↗
PubMed1989

Dermorphin analogues carrying an increased positive net charge in their "message" domain display extremely high mu opioid receptor selectivity.

According to the membrane compartment concept the receptor specificity of ligands is based not only on ligand-receptor complementarity but also on specific ligand-membrane interactions. Elaboration of this concept for opioid peptide-receptor interactions had led to the assumption that mu- and delta-receptors are located in anionic and cationic membrane compartments, respectively, and to the prediction that positively charged opioid receptor ligands should display mu-receptor selectivity. To assess the validity of this model, we synthesized a series of dermorphin analogues carrying a net positive charge and tested them in mu- and delta-receptor representative binding assays and bioassays. Some but not all of the prepared compounds showed the receptor-selectivity profile expected on the basis of the membrane compartment concept. In particular, gradual augmentation of the positive charge from 1+ to 3+ in a series of dermorphin-(1-4) tetrapeptide analogues produced an enhancement of mu-receptor affinity and a progressive decrease in delta-receptor affinity, resulting in increasingly higher mu-receptor selectivity. The most selective compound was [D-Arg2,Lys4]dermorphin-(1-4)-amide (DALDA), showing a selectivity ratio (Ki delta/Ki mu = 11,400) more than 10 times higher than that of DAGO (Ki delta /Ki mu = 1050) and, thus, displaying unprecedented mu-receptor specificity. Because of its high positive charge (3+), DALDA may be particularly useful as a very specific agonist for studying peripheral mu-receptor interactions.

Animals↗
PubMed1988

A mutation in the 530 loop of Escherichia coli 16S ribosomal RNA causes resistance to streptomycin.

Oligonucleotide-directed mutagenesis was used to introduce an A to C transversion at position 523 in the 16S ribosomal RNA gene of Escherichia coli rrnB operon cloned in plasmid pKK3535. E. coli cells transformed with the mutated plasmid were resistant to streptomycin. The mutated ribosomes isolated from these cells were not stimulated by streptomycin to misread the message in a poly(U)-directed assay. They were also restrictive to the stimulation of misreading by other error-promoting related aminoglycoside antibiotics such as neomycin, kanamycin or gentamicin, which do not compete for the streptomycin binding site. The 530 loop where the mutation in the 16S rRNA is located has been mapped at the external surface of the 30S subunit, and is therefore distal from the streptomycin binding site at the subunit interface. Our results support the conclusion that the mutation at position 523 in the 16S rRNA does not interfere with the binding of streptomycin, but prevents the drug from inducing conformational changes in the 530 loop which account for its miscoding effect. Since this effect primarily results from a perturbation of the translational proofreading control, our results also provide evidence that the 530 loop of the 16S rRNA is involved in this accuracy control.

Drug Resistance, Microbial↗
PubMed1988

The chloroplast genome of the green alga Chlamydomonas moewusii: localization of protein-coding genes and transcriptionally active regions.

We have recently reported the map positions of the ribosomal RNA genes and 6 protein-coding genes on the 292 kbp Chlamydomonas moewusii chloroplast genome. In the present study we localized 12 additional protein-coding genes on this green algal genome as well as 5 of these genes on the 196 kbp C. reinhardtii chloroplast genome. The gene mapping data agree with previous reports indicating that these two algal genomes differ tremendously in their global sequence organization and bear little similarity to their land plant counterparts. Among the 18 protein-coding genes examined, atpA and atpF of C. moewusii constitute the unique set of closely linked genes which is shared with land plant chloroplast genomes. The important gene order differences between the C. moewusii and C. reinhardtii chloroplast DNAs led us to speculate that gene grouping into transcription units is less prevalent in green algal chloroplast genomes than in their land plant homologs. To gain an insight into the transcriptional organization of the C. moewusii chloroplast genome, we probed Northern blots of total cellular RNA with clones covering 85% of this genome. Most chloroplast DNA regions were found to produce simple transcript patterns with RNAs less than 3.5 kb in size. This may be interpreted as indicating that Chlamydomonas chloroplast genes are transcribed mainly as monocistronic RNAs or that the RNAs observed are processed forms of unstable polycistronic transcripts.

Biological Evolution↗
PubMed1988

Mapping of chloroplast mutations conferring resistance to antibiotics in Chlamydomonas: evidence for a novel site of streptomycin resistance in the small subunit rRNA.

A major obstacle to our understanding of the mechanisms governing the inheritance, recombination and segregation of chloroplast genes in Chlamydomonas is that the majority of antibiotic resistance mutations that have been used to gain insights into such mechanisms have not been physically localized on the chloroplast genome. We report here the physical mapping of two chloroplast antibiotic resistance mutations: one conferring cross-resistance to erythromycin and spiramycin in Chlamydomonas moewusii (er-nM1) and the other conferring resistance to streptomycin in the interfertile species C. eugametos (sr-2). The er-nM1 mutation results from a C to G transversion at a well-known site of macrolide resistance within the peptidyl transferase loop region of the large subunit rRNA gene. This locus, designated rib-2 in yeast mitochondrial DNA, corresponds to residue C-2611 in the 23 S rRNA of Escherichia coli. The sr-2 locus maps within the small subunit (SSU) rRNA gene at a site that has not been described previously. The mutation results from an A to C transversion at a position equivalent to residue A-523 in the E. coli 16 S rRNA. Although this region of the E. coli SSU rRNA has no binding affinity for streptomycin, it binds to ribosomal protein S4, a protein that has long been associated with the response of bacterial cells to this antibiotic. We propose that the sr-2 mutation indirectly affects the nearest streptomycin binding site through an altered interaction between a ribosomal protein and the SSU rRNA.

Base Sequence↗
PubMed1988

Cloning and sequencing of a cDNA encoding the small subunit precursor of ribulose-1,5-bisphosphate carboxylase from Chlamydomonas moewusii. Evolution of RUBISCO SS polypeptide.

We have isolated and characterized a full-length cDNA clone encoding the precursor of the small subunit (pSU) of ribulose-1,5-bisphosphate carboxylase/oxygenase (RUBISCO) from the green alga, Chlamydomonas moewusii. Comparison with the C. reinhardtii rbcS1 gene sequence reveals that both small subunit (SS) coding regions are 75% homologous and that their predicted mature polypeptide chains are each composed of 140 amino acids. In contrast, their transit peptides appear to be divergent. We also show that transcription of the C. moewusii rbcS gene(s) which generates a 1,230 and a 930 base mRNA species are light-stimulated/or accumulated during the light period of the cell cycle. Finally, the SS polypeptide sequences of fifteen different photosynthetic organisms are compared; this analysis reveals at least five well-conserved polypeptide domains.

Amino Acid Sequence↗
PubMed1988

Interindividual and intraindividual variability in acetylation: characterization with caffeine.

The degree of interindividual and intraindividual variability in acetylator activity was investigated with caffeine used as a probe of enzyme activity. Acetylator phenotype and relative N-acetyltransferase activity were estimated in 46 subjects by measuring the urinary ratio of two metabolites, AFMU/1-MX, after a single 300 mg oral dose of caffeine on five separate occasions. Thirty homozygous slow (rr) and 15 heterozygous rapid (Rr) acetylators were identified. The degree of interindividual variability in acetylator activity was observed to be a mean of 32% (range 27% to 36%) and 20% (range 11% to 29%) in the rr and Rr groups, respectively. The mean intraindividual variation on repetitive measurement was 19% (range 6% to 49%) in the rr and 14% (range 7% to 24%) in the Rr acetylator group. Four subjects had apparent changes in acetylator activity with time such that they were unable to be assigned to any one acetylator group. Two of these four subjects exhibited apparent homozygous rapid acetylator activity intermittently during the 5-week trial. This variability may explain, in part, some of the high degree of patient variability observed in the toxicity, efficacy, and drug-related disease associated with acetylated drugs and environmental toxins.

Acetylation↗
PubMed1988

Synthesis and biological activity of [Leu5]enkephalin derivatives containing D-glucose.

The synthesis of some [Leu5]enkephalin derivatives is described in which D-glucose has been linked to the opioid pentapeptide through the ester bond involving the carboxyl function at the C-terminal with C-1 or C-6 of the D-glucopyranose moiety. Enkephalin derivatives were assayed for opioid activity and found to be full agonists in bioassays based on inhibition of electrically evoked contractions of the guinea pig ileum (GPI) and of the mouse vas deferens (MVD). The obtained results suggest that the opioid activity of the tested glucoconjugates depend upon the ester bond position in the molecule. Whereas 1-O conjugate 5 was somewhat more potent than [Leu5]enkephalin in the GPI assay, the 6-O conjugates, with the exception of 1-O-benzyl derivative 11, were considerably less potent. All enkephalin derivatives were delta-receptor selective; in particular, the acetylated analog 8 was three times more delta-receptor selective than [Leu5]enkephalin.

Animals↗