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Biomedical subjects

F Ramirez

Publications and source records attributed to F Ramirez.

At least 145 records · Page 8Linked to original sources
PubMed1984

Lateral interaction of cholesterol in diacylphosphatidylcholesterol bilayers.

Thermotropic phase-transition properties of the aqueous dispersions of several diacylphosphatidylcholesterol (DRCh) analogs are examined. The aqueous dispersions of their calcium salts exhibit characteristic endothermic thermotropic transitions due to a change in the conformation of acyl chains. These dispersions consist of osmotically intact liposomes that trap ions, and at the transition temperature there is anomalous increase in the ion leakage. Wide-angle electron diffraction studies of DPCh . Ca monolayers also exhibit a transition from a sharp 4.25 A band to a broad one centering at 4.7 A, reflecting an order-disorder transition in the acyl chains. The long-range order in the organization of acyl chains of DRCh molecules could arise from intermolecular interactions between the cholesterol moieties to form a functional dimer, and such dimers are apparently cross-linked by Ca2+ to form a long-range interacting lattice of acyl chains. Evidence for this model is adduced from the fluorescence properties of the dispersions of dimyristoylphosphatidylcholesta-5,7,9-trienol. The phase-transition properties of DRCh are an ideal illustration of calcium-induced isothermal phase transition.

Cholestanes↗
PubMed1984

Evidence from oxygen exchange studies that the two heads of myosin are functionally different.

Recent studies of oxygen exchange have shown that there are two normal pathways for the hydrolysis of MgATP by myosin in the presence of actin, each producing Pi at the same rate. These two apparent pathways for actin-activated hydrolysis differ greatly in the extent of oxygen exchange they support. This is revealed by an analysis of the distribution of [18O]Pi species produced by the hydrolysis of [gamma-18O]ATP. We have extended these studies to certain abnormal substrates, using Mn2+ in place of Mg2+, and dATP or ITP in place of ATP. The results, together with past findings, lead to the proposal that the two heads of myosin are functionally different. One of these (Head 1) is able to reversibly cleave bound MgATP and thereby support oxygen exchange while it is free of actin; the other (Head 2) cannot cleave bound MgATP at its active site while free of actin. However, in the presence of actin, both Head 1 and Head 2 cleave bound MgATP, support some oxygen exchange, and produce Pi at the same rapid actin-activated rate. Apparently, MgATP is positioned differently on the two heads when they are free of actin, with Mg2+ and the 6-amino group of ATP playing an important role in the orientation. This proposed difference between the heads could serve to make Head 1 react first with the actin filament, followed by Head 2. Thus, in muscle, the two heads on a myosin cross-bridge would interact with an actin filament and exert their pull in a fixed sequence.

Actins↗
PubMed1984

Amine fluorescamine compounds inhibit oxidative phosphorylation in rat liver mitochondria.

The reaction of fluorescamine with ammonia, benzylamine, o,p-dimethylbenzylamine, 2-phenylethylamine, p-aminobenzoic acid, and the mycosamine-containing macrolide antibiotic, amphotericin B, yield compounds which induce significant effects on mitochondrial activities. From their effects on energy-yielding processes which lead to transmembranous proton movements, the compounds may be divided into three classes. While all modifiers significantly inhibit proton movement induced by both ATP hydrolysis and electron transfer in mitochondria, their influence on the primary energy yielding steps are quite different. Class I modifiers, e.g., the compound made from amphotericin B, inhibit electron transfer but have no effect on the Pi release associated with ATP hydrolysis. Class II modifiers, e.g., the compound made from benzylamine, inhibit respiration but stimulate Pi release. Class III modifiers, e.g., the compound made from p-aminobenzoic acid, on the other hand, only slightly increase Pi release but have no effect on redox reactions. These and other effects of the modifiers are taken to mean that the proton movements and their associated energy-yielding processes are only linked indirectly. The effects of the modifiers on State 3 mitochondrial activities were also investigated. Although all the modifiers decrease the rates of both State 3 respiration and its coupled ATP synthesis, the efficiency of energy conversion measured by the P/O ratio remains unaltered.

Adenosine Triphosphatases↗
PubMed1984

Molecular heterogeneity in the mild autosomal dominant forms of osteogenesis imperfecta.

Mild osteogenesis imperfecta (OI type I and OI type IV) is characterized by postnatal onset of fractures, absence of skeletal deformity, presenile hearing loss with or without blue sclerae, and dentinogenesis imperfecta. Using one common DNA polymorphism associated with the pro alpha 2(I) human collagen gene, we found genetic heterogeneity in this disorder. In three families, the OI phenotype segregated independently of the DNA polymorphism, whereas in one family, the OI phenotype cosegregated with a DNA polymorphism in a manner suggesting linkage. Use of DNA polymorphisms associated with both type I procollagen genes should provide a tool to unravel the molecular heterogeneity of various heritable disorders of the connective tissue.

DNA Restriction Enzymes↗
PubMed1983

Nucleotide sequences of complementary deoxyribonucleic acids for the pro alpha 1 chain of human type I procollagen. Statistical evaluation of structures that are conserved during evolution.

Nucleotide sequences were determined for two cloned cDNAs encoding for over three-fourths of the pro alpha 1 (I) chain of type I procollagen from man. Comparison with previously published data on amino acid sequences of the alpha 1 (I) chain of type I collagen made it possible to examine mutations in the transcribed products of the gene which have occurred during the evolution of man, calf, rat, mouse, and chick. Comparison of the nucleotide sequences with the corresponding sequences of cDNAs from chick [Fuller, F., & Boedtker, H. (1981) Biochemistry 20, 996] and with cDNAs for the pro alpha 2(I) chain from man [Bernard, M.P., Myers, J. C., Chu, M.-L., Ramirez, F., Eikenberry, E. F., & Prockop, D. J. (1983) Biochemistry 22, 1139] demonstrated that selective pressure during evolution for 250 million or more years acted more strongly on the structure of the pro alpha 1 (I) chain than on the pro alpha 2(I) chain. To improve the reliability of the comparison, the nucleotide sequences were examined with a modification of previous procedures for evaluating mutations in replacement sites and silent sites. The corrected divergence for replacement sites between the alpha 1 (I) chains was 6 +/- 0.8% whereas it was 15 +/- 1.9% for the alpha 2(I) chains. The C-propeptide domain of the pro alpha 1 (I) chain was also highly conserved with a corrected divergence at replacement sites of 5 +/- 0.9%, a value that was not distinguishable from the value previously found for the C-propeptide of the pro alpha 2(I) chain. Therefore, a large part of the structure of both C-propeptides appears to be under selective pressure. Inspection of changes in the C-propeptide of the pro alpha 1 (I) chain suggested that there was a highly conserved region around the carbohydrate attachment site similar to the highly conserved region of 37 amino acids previously found in the C-propeptide of the pro alpha 2(I) chain. Two statistical tests, however, were unable to confirm nonrandom distribution of changes in the C-propeptide of the pro alpha 1(I) chain. The same tests established the presence of a nonrandom distribution in nucleotide changes of the C-propeptide of the pro alpha 2(I) chain. The 3'-noncoding region of the cDNA for pro alpha 1(I) of human type I procollagen showed no homology with the same region in the chick.(ABSTRACT TRUNCATED AT 400 WORDS)

Amino Acid Sequence↗
PubMed1983

Oxygen-exchange studies on the pathways for magnesium adenosine 5'-triphosphate hydrolysis by actomyosin.

At an intermediate stage in the hydrolysis of magnesium adenosine 5'-phosphate (MgATP) by myosin or actomyosin, there is an exchange of oxygen between water and the P gamma group of enzyme-bound nucleotide. Starting with [P gamma-18O]ATP as substrate, the exchange is revealed in the [18O]Pi species that are ultimately released as product into the reaction medium. An analysis of the distribution of these labeled Pi species, which contain 3, 2, 1, or none of the 18O atoms originally on the P gamma of ATP, is used to probe intermediate stages of the hydrolytic mechanism. In recent years, studies of this kind by several groups have shown that more than one pathway of hydrolysis operates. The work reported here demonstrates that two of these pathways are spurious; one is a "nonexchanging MgATPase" that is present in fresh myosin preparations; the other is an induced slow exchange that develops in myosin during storage (-20 degrees C) and subsequent aging (4 degrees C). However, after correction for these artifacts, two normal pathways for actomyosin hydrolysis remain. These normal pathways differ in the mode of interaction between actin and myosin in the course of hydrolysis; one is the Lymn-Taylor pathway where oxygen exchange occurs at a stage when actin and myosin are dissociated; the other is a pathway in which actin and myosin are associated during oxygen exchange. Each of these two pathways contributes an equal amount of Pi to the product pool. Thus, on average, each myosin head uses each of these pathways half the time. The findings suggest, e.g., that during contraction, myosin can dissociate from the actin filament only during every other cycle of MgATP hydrolysis or that only half the heads, at any one time, can exchange oxygen while free of the actin filament.

Actomyosin↗
PubMed1983

Analysis of the 3' end of the human pro-alpha 2(I) collagen gene. Utilization of multiple polyadenylation sites in cultured fibroblasts.

Three overlapping genomic clones covering 28 kilobases of the human pro-alpha 2(I) collagen gene have been isolated from a lambda phage library. The analysis of 12 introns and 12 exons in the 3' end region has shown that the human gene has a structure remarkably similar to that reported for the homologous chicken gene. One large intron, in the alpha-chain domain, contains an AluI sequence flanked by short direct repeats; a second AluI sequence is present 4 kilobases downstream from the termination codon. The analysis of the exon coding for the 3'-untranslated region has revealed that the pro-alpha 2(I) collagen gene transcribes at least four different mRNAs in cultured fibroblasts. The colinearity and exact location of the termini of these transcripts was determined by Northern blots, R-looping analysis, S1 protection, and DNA sequencing. The ends of two transcripts are closely preceded by the canonical polyadenylation signal (AAUAAA), whereas two of its variations (AUUAAA and AUUAA) precede the ends of the other two transcripts.

Base Sequence↗
PubMed1983

Structure of a cDNA for the pro alpha 2 chain of human type I procollagen. Comparison with chick cDNA for pro alpha 2(I) identifies structurally conserved features of the protein and the gene.

Nucleotide sequences were determined for cloned cDNAs encoding for more than half of the pro alpha 2 chain of type I procollagen from man. Comparisons with previously published data on homologous cDNAs from chick embryos made it possible to examine evolution of the gene in two species which have diverged for 250-300 million years. The amino acid sequence of the alpha-chain domain supported previous indications that there is a strong selective pressure to maintain glycine as every third amino acid and to maintain a prescribed distribution of charged amino acids. However, there is little apparent selective pressure on other amino acids. The amino acid sequence of the C-propeptide domain showed less divergence than the alpha-chain domain. The 5' end or N terminus of the human C-propeptide, however, contained an insert of 12 bases coding for 4 amino acids not found in the chick C-propeptide. About 100 amino acid residues from the N terminus, two residues found in the chick sequence were missing from the human. In the second half of the C-propeptide, there was complete conservation of a 37 amino acid sequence and conservation of 50 out of 51 amino acids in the same region, an observation which suggested that the region serves some special purpose such as directing the association of one pro alpha 2(I) C-propeptide with two pro alpha 1(I) C-propeptides so as to produce the heteropolymeric structure of type I procollagen. In addition, comparison of human and chick DNAs for pro alpha 2(I) revealed three different classes of conservation of nucleotide sequence which have no apparent effect on the structure of the protein: a preference for U on the third base position of codons for glycine, proline, and alanine; a high degree of nucleotide conservation in the 51 amino acid highly conserved region of the C-propeptide; a high degree of nucleotide conservation in the 3'-noncoding region. These three classes of nucleotide conservation may reflect unusual features of collagen genes, such as their high GC content or their highly repetitive coding sequences.

Amino Acid Sequence↗
PubMed1983

Restriction fragment length polymorphism associated with the pro alpha 2(I) gene of human type I procollagen. Application to a family with an autosomal dominant form of osteogenesis imperfecta.

One cloned complementary DNA and one genomic subclone were used to detect restriction fragment length polymorphism associated with the pro alpha 2(I) gene for human type I procollagen. The restriction fragments obtained from examination of 30-122 chromosomes confirmed previous indications that the pro alpha 2(I) gene is found in a single copy in the human haploid genome. One highly polymorphic site was detected with EcoRI in the 5'-half of the gene. The restriction site polymorphism at the site had an allelic frequency of 0.38, and it generated two fragments of 10.5 and 3.5 kilobase in homozygous individuals. The restriction fragment length polymorphism generated at the EcoRI site was used to study affected and non-affected individuals in four generations of a family with an autosomal dominant form of osteogenesis imperfecta. The data demonstrated a linkage of the phenotype to a pro alpha 2(I) allele with a lod score of 2.41 at a recombination fraction (theta) of 0. The data therefore provided presumptive evidence that osteogenesis imperfecta in this family is caused by a mutation in the pro alpha 2(I) gene or some contiguous region of the genome. The relatively high frequency of polymorphism at the EcoRI site makes it useful for studying a broad range of genetic disorders in which mutations in type I procollagen are suspected. In addition, the polymorphic site should provide useful markers for linkage studies with other loci located on human chromosome 7.

DNA Restriction Enzymes↗
PubMed1982

Cloning and characterization of five overlapping cDNAs specific for the human pro alpha 1(I) collagen chain.

We report the cloning of five overlapping cDNAs bearing sequences specific for the human pro alpha 1(I) collagen chain. Poly-A RNA enriched for collagen sequences was purified from normal human fibroblasts and used as template to synthesize double stranded cDNA. The cDNA was inserted into the Eco RI site of pBR 322 by blunt-ending and dG:dC tailing. The clones were screened by colony hybridization using the original RNA population and the resulting five positive clones subjected to restriction endonuclease mapping analysis and DNA sequencing. These overlapping clones cover from residue 247 in the alpha chain to part of the 3' end untranslated region of the pro alpha 1(I) mRNA for a total of 3400 nucleotides.

Amino Acid Sequence↗
PubMed1982

Inhibition of oxygen exchange by chemical modifiers at the sulfhydryl 1 or reactive lysine residue of myosin. Changing the rates of intermediate enzymatic reactions by modifying the course of reaction-linked conformational changes.

During the hydrolysis of MgATP by myosin, there is an extensive exchange of oxygen between water and the terminal phosphate group of bound nucleotide, which results from a repeated cycle of hydrolytic cleavage and its reversal. An analysis of the distribution of [18O]Pi species from the hydrolysis of [gamma-18O]ATP gives an estimate for the apparent rate constant [k-3(app)] of reverse cleavage, which is the rate-limiting step of the exchange cycle. We have performed this kind of analysis with two different forms of modified myosin, containing either N-ethylmaleimide at the sulfhydryl 1 group or trinitrophenyl at the reactive lysine residue. Although these modifications of the protein are chemically different, and the sulfhydryl 1 group and the reactive lysine residue are far apart in the primary chain of the myosin head, the two modifications caused a similar marked inhibition of oxygen exchange. This effect resulted from: 1) a decrease in the time available for the exchange cycle due to a 3-10-fold increase in the turnover rate of hydrolysis and 2) a reduction in k-3 (app) from 2-5 s-1 to 0.2-0.5 s-1. It is proposed that modifications of this type influence enzymatic activity by altering the course of "reaction-linked conformational changes," i.e. those changes in protein structure that are coupled to the catalytic mechanism but are not part of the substrate-binding site proper. It is also suggested that certain naturally occurring protein modifications, e.g. methylation of histidine or lysine residues, could regulate function by operation of this mechanism.

Adenosine Triphosphate↗
PubMed1982

Idiopathic membranous nephropathy in children.

To determine the prognosis in children with membranous nephropathy, we reviewed the clinicopathologic features and outcome of 22 patients, 11 male and 11 female, age 11 months to 19.9 years (mean 12.0 years). Patients had biopsies within six months of onset of symptoms and were divided into two groups according to the biopsy findings: group 1 (Stages I and II) and group 2 (Stages III and IV). Follow-up time was identical in both groups (mean 4.8 years). The nephrotic syndrome was present in 13 of 16 patients in group 1 and in all six patients in group 2. In group 1, eight of the 16 patients had repeat biopsies one to 11 years (mean three years) after onset. Of these, two progressed to Stages III and IV, whereas progression to renal insufficiency occurred in only one. In group 2, two of the six patients had repeat biopsies two years after onset and remained in the same group; progression to renal insufficiency occurred in five of the six patients. The difference in progression to renal insufficiency between the two groups is significant (P = 0.001). The stage of glomerular lesion at the time of onset seems to be a factor in predicting the prognosis of membranous nephropathy in pediatric patients.

Adolescent↗
PubMed1982

Human type I procollagen genes are located on different chromosomes.

A recombinant plasmid containing sequences complementary to human pro-alpha l(I) collagen mRNA was used for the chromosomal assignment of the pro-alpha l(I) collagen gene. Restriction endonuclease analysis of DNA from mouse-human and Chinese hamster-human somatic cell hybrids revealed cosegregation with human chromosome 17. Hybrids containing derivative chromosomes with a t(2;17)(q14;q21) translocation showed cosegregation of the pro-alpha l(I) gene with the segment 17q21 leads to qter. In situ hybridization on human metaphasic chromosomes confirmed this conclusion.

Amino Acid Sequence↗
PubMed1982

Neonatal pemphigus vulgaris: role of passively transferred pemphigus antibodies.

A neonate with oral lesions of pemphigus vulgaris is described. The diagnosis was confirmed by cytological examination of lesions, positive intercellular IgG staining of the acantholytic cells, and presence of pemphigus antibodies in his serum. At the time of delivery, the child's mother had active pemphigus vulgaris, and it is suggested that this was a case of passively transferred disease.

Adolescent↗
PubMed1982

Effects of borohydride-treated oligomycins on processes of energy transduction in mitochondria.

Sodium borohydride in ethanol solution under mild conditions brings about the stepwise reduction of the 7-keto and the 11-keto groups of rutamycin and the oligomycins to the corresponding hydroxyl groups without further alterations of the macrocyclic lactone structure or other features of the molecule. The reduced compounds, as well as the parent antibiotics, inhibit the ADP-dependent (state 3) respiration, and the Pi formation and proton extrusion that are linked to ATP hydrolysis, but have no effect on other respiration-linked activities in intact rat liver mitochondria. Analogous inhibitory effects of borohydride-treated antibiotics are also observed in rat-liver submitochondrial particles. The reduced compounds are less potent inhibitors than the parent antibiotics. The reduced compounds are more efficient as inhibitors of Pi formation stimulated by conventional uncouplers (e.g. 2,4-dinitrophenol), than of Pi formation stimulated by certain amine-fluorescamine modifiers (e.g.) the benzylamine-fluorescamine compound. In contrast, the parent antibiotics are unable to discriminate between uncoupler-stimulated and modifier-stimulated Pi formation. It is suggested that rutamycin and the oligomycins bind to H+-ATPase as a result of hydrogen bonding to, at least, the 7-keto and/or the 11-keto groups of the antibiotics. When these keto groups are reduced to hydroxyl groups the hydrogen-bonding is less efficient due to the pronounced directional characteristic of hydrogen-bonding to keto groups.

Adenosine Triphosphatases↗
PubMed1982

Effects of hemodialysis and dimercaprol in acute dichromate poisoning.

A 22-month-old infant died after ingesting sodium dichromate his father had brought from work. Treatment included folic acid and dimercaprol administration, hemodialysis, and exchange transfusion. To evaluate this treatment, four dogs were hemodialyzed after receiving intravenous sodium dichromate: their dialyzer chromate clearance was similar to their renal chromate clearance and their dialyzer chromate clearance was not significantly different before or after dimercaprol administration. This and other cases in the literature indicate that although chromate poisoning is often fatal, supportive care, forced diuresis, and chelating agents may be helpful. Hemodialysis may be required if renal failure occurs. Awareness of toxicity and prevention remain the most important approaches.

Acute Disease↗