Biomedical subjects

V Ramakrishnan

Publications and source records attributed to V Ramakrishnan.

At least 163 records · Page 9Linked to original sources

PubMed1994

Insomnia in Vietnam era veteran twins: influence of genes and combat experience.

Genetic and environmental influences on insomnia were studied in 2,825 pairs of Vietnam era veteran male twins. The self-reported sleep problems studied included trouble falling asleep, trouble staying asleep, waking often, waking tired and a composite sleep scale. Twin correlations for each of the sleep problems were larger in monozygotic than in dizygotic pairs, with heritability estimates ranging from 0.21 to 0.42. There was no effect of common familial environment. Phenotypic correlations for combat experience and sleep problems were small, ranging from 0.00 to 0.09, with no differences seen in monozygotic and dizygotic twins. When the effects of genes and combat exposure were evaluated simultaneously, there was a significant genetic contribution to all sleep measures, but combat exposure was significantly associated only with overall sleep quality, waking often and having trouble staying asleep.

Adult↗

PubMed1994

The anion-binding exosite is critical for the high affinity binding of thrombin to the human thrombin receptor.

The thrombin receptor has been shown to be a novel member of the family of G-protein coupled receptors (Vu, T.-K. H., Hung, D.T., Wheaton, V.I., and Coughlin, S.R. (1991) Cell 64, 1057-1068). This receptor appears to be activated through a thrombin-mediated proteolytic mechanism which exposes a "tethered ligand" responsible for receptor activation. In order to investigate the initial interactions of thrombin with this receptor, we have constructed cell lines which express high levels of the human thrombin receptor and studied the binding of various forms of thrombin to the cell surface. Analysis of transfected cells with thrombin receptor monoclonal antibodies identified a particular cell line (clone #5-18) which displayed > 150,000 thrombin receptors per cell. Clone #5-18 appeared to express functional receptors since treatment with thrombin resulted in both a 15-20 fold increase of cytoplasmic phosphoinositide levels and a comparable shift in the EC50 of thrombin-mediated calcium mobilization when compared to non-transfected CHO cells. Binding of 125I-alpha-thrombin to clone #5-18 did not reach equilibrium at 37 degrees C. However, direct binding studies of 125I-alpha-, 125I-diisopropylphospho (DIP)-alpha-, and 125I-beta-thrombin to clone #5-18 demonstrated that binding at 4 degrees C was saturable and reversible for each ligand. Analysis of the binding data revealed Kd's of 0.8 nM, 0.7 nM and 9.7 nM for 125I-alpha-, 125I-DIP-alpha- and 125I-beta-thrombin respectively. Association of 125I-alpha-, DIP-alpha, and beta-thrombin could be competed by unlabelled alpha- and DIP-alpha-thrombin. Unlabelled beta-thrombin, which has a modified anion-binding exosite, was a poor competitor for 125I-alpha- and 125I-DIP-alpha-thrombin, but did compete for 125I-beta-thrombin. In addition, the hirudin54-65 peptide competed at submicromolar concentrations for the binding of alpha- and DIP-alpha-thrombin, but not for beta-thrombin. This peptide binds specifically at the anion-binding exosite of alpha-thrombin and has been shown to have a lower affinity for beta-thrombin. These results demonstrate directly a high affinity interaction between thrombin and its receptor, and suggest that an important component is the high affinity association of the thrombin receptor with the anion-binding exosite of thrombin.

Amino Acid Sequence↗

PubMed1994

Ultrasonographic growth and Doppler hemodynamic evaluation of fetuses of obese women.

All fetuses benefit from ultrasonographic estimation of gestational age and evaluation of growth patterns. Monitoring the pregnancies of obese women is perceived as more difficult than monitoring those of nonobese women. The aim of this study was to determine if maternal obesity affects the growth and Doppler resistance indices (RI) of the fetus. Twenty-eight women with a preconception weight > 90.7 kg underwent obstetric ultrasonographic evaluations from the 20th week of gestation. Their ultrasonographic data were compared with those of controls. Ten of the obese women developed gestational diabetes and had lower umbilical artery RIs for a given gestational age (P < .0001) than did those obese women without other medical complications, those with medically controlled pregnancy-induced hypertension or those from the control population. The relation between fetal unit weight and umbilical artery RIs was established. The pattern of RI changes was similar in all groups when estimated fetal weight instead of gestational age was used as the covariant. Fetal growth and Doppler velocimetry can be monitored adequately in obese women. Gestational diabetes significantly influences the pattern of fetal growth and the impedance to flow in the umbilical artery.

Adult↗

PubMed1993

Ribosomal protein L6: structural evidence of gene duplication from a primitive RNA binding protein.

In all cells, protein synthesis is coordinated by the ribosome, a large ribonucleoprotein particle that is composed of > 50 distinct protein molecules and several large RNA molecules. Here we present the crystal structure of ribosomal protein L6 from the thermophilic bacterium Bacillus stearothermophilus solved at 2.6 A resolution. L6 contains two domains with almost identical folds, implying that it was created by an ancient gene duplication event. The surface of the molecule displays several likely sites of interaction with other components of the ribosome. The RNA binding sites appear to be localized in the C-terminal domain whereas the N-terminal domain contains the potential sites for protein-protein interactions. The domain structure is homologous with several other ribosomal proteins and to a large family of eukaryotic RNA binding proteins.

Animals↗

PubMed1993

Ribosomal protein S17: characterization of the three-dimensional structure by 1H and 15N NMR.

The structure of ribosomal protein S17 from Bacillus stearothermophilus was investigated by two-dimensional homonuclear and heteronuclear magnetic resonance spectroscopy. The 1H and 15N chemical shift assignments are largely complete, and a preliminary structural characterization is presented. The protein consists of five beta-strands that form a single antiparallel beta-sheet with Greek-key topology. The beta-strands are connected by several extended loops, and two of these contain residue types that are frequently seen in the RNA-binding sites of proteins. Additionally, two point mutations that affect antibiotic resistance, translational fidelity, and ribosome assembly are located in these two regions of the protein. Since these potential RNA-binding sites are distributed over a large surface of the protein, it appears that the molecule may interact with several regions of 16S rRNA.

Amino Acid Sequence↗

PubMed1993

Homo- and heteronuclear two-dimensional NMR studies of the globular domain of histone H1: sequential assignment and secondary structure.

A recombinant 75 amino acid polypeptide corresponding to the globular domain of the chicken histone H1 (GH1) has been studied by 1H homonuclear and 1H-15N heteronuclear 2D NMR spectroscopy. Sequential assignment of the backbone and beta-proton resonances has enabled us to determine the secondary structure of GH1. It was found to consist of three helical regions (T7-S17, L25-Y37, E40-K56) and probably a beta-hairpin (L59-L73). This structure is similar to the structure of the globular domain of histone H5 (GH5) obtained both by NMR spectroscopy [Zarbock et al. (1986) Proc. Natl. Acad. Sci. U.S.A. 83, 7628-7632; Clore et al. (1987) EMBO J. 6, 1833-1842] and by X-ray crystallography [Ramakrishnan et al. (1993) Nature 362, 219-223]. The beta-hairpin as suggested for GH1 is also present in the X-ray structure of GH5 but has not been reported for the NMR structure of GH5.

Amino Acid Sequence↗

PubMed1993

A strategy for assembling samples of adult twin pairs in the United States.

In this paper we develop a methodology for the identification of large numbers of U.S. adult twin pairs. Data for this study derive from the U.S. Department of Defense and the Vietnam Era Twin (VET) Registry. The Department of Defense identified potential male twins (n = 10,002) using a computerized record linkage algorithm based on the same last name, same date of birth, and the same first five digits of the Social Security number. Twinship was confirmed by comparison with the Vietnam Era Twin Registry. We developed a logistic regression model that predicts the probability that a paired record identifies twins based on the absolute difference in the last four digits in the Social Security number, the age of issuance of the Social Security number, and the frequency of occurrence of the last name. We used the estimated coefficients derived from this regression model to assign predicted probabilities of being a twin to each matched record. There is a close correspondence between the observed and expected number of twins when evaluated across deciles of predicted probabilities of being a twin; the value of the Harrell's c index (c = 0.68 +/- 0.0004) indicates the overall predictive accuracy of the regression equation. The results from this study demonstrate the feasibility of identifying adult male-male twin pairs from any large computerized database that contains name, date of birth and Social Security number. However, the selection criteria used in the creation of the computer database must be clearly specified to avoid constructing a biased sample of twins.

Adult↗

PubMed1993

Crystal structure of globular domain of histone H5 and its implications for nucleosome binding.

The structure of GH5, the globular domain of the linker histone H5, has been solved to 2.5 A resolution by multiwavelength anomalous diffraction on crystals of the selenomethionyl protein. The structure shows a striking similarity to the DNA-binding domain of the catabolite gene activator protein CAP, thereby providing a possible model for the binding of GH5 to DNA.

Amino Acid Sequence↗

PubMed1993

Kinetics of CD11b expression on neutrophils isolated from subjects with healthy gingivae and patients with advanced periodontitis.

The adhesion molecule, CD11b/CD18, plays a role in host defense against bacterial infections owing to its involvement in cell migration and phagocytosis. We examined the kinetics of CD11b expression on neutrophils isolated from peripheral blood (PB-PMNs) and gingival crevicular fluid (CF-PMNs) in subjects with healthy gingivae and in patients with advanced periodontitis in order to assess if CF-PMNs respond differently in regard to CD11b expression from PB-PMNs isolated from healthy or periodontitis subjects. CF-PMNs or formylmethionylleucylphenylalanine (fMLP)-stimulated PB-PMNs were incubated in buffer with or without divalent cations for 60, 90, or 110 minutes. Plasma membrane (surface) CD11b as measured by flow cytometry was found to be reduced with time when cells (PB-PMNs or CF-PMNs) from either controls or patients were treated with calcium- and magnesium-free buffer. However, when CF-PMNs were treated with buffer containing Ca++/Mg++, surface CD11b expression increased on cells from both controls and patients. The reduction in CD11b expression on PB-PMNs is significantly greater than that measured on CF-PMNs over time. Therefore, CF-PMNs respond differently from PB-PMNs which have been artificially stimulated with fMLP in regard to kinetics of CD11b expression on the plasma membrane.

Adult↗

PubMed1993

A novel monoclonal antibody dependent on domain 5 of the platelet-derived growth factor beta receptor inhibits ligand binding and receptor activation.

Platelet derived growth factor (PDGF) induces activation of the protein tyrosine kinase domain of the PDGF receptor, resulting in receptor dimerization and the initiation of mitogenesis in responsive cells. In order to identify domains of the receptor involved in these processes, a panel of monoclonal antibodies (MAbs) against the extracellular region of the human PDGF receptor was developed and screened to identify which of these specifically block PDGF binding. One of these, MAb 2A1E2, binds PDGF beta receptor with high affinity and blocks PDGF BB binding in a whole cell binding assay with an IC 50 of 0.1 nM. Inhibition of binding results in the inhibition of ligand-induced receptor phosphorylation, dimerization and mitogenesis in cells expressing the PDGF beta receptor. MAb 2A1E2 has been mapped to the fifth Ig domain of the PDGF beta receptor, implying that this domain is important for ligand binding, dimerization and/or activation. The potency of MAb 2A1E2 for inhibiting PDGF BB binding indicates that this antibody is ideally suited to identify and characterize PDGF BB-induced biological responses.

Animals↗

PubMed1992

The structure of ribosomal protein S5 reveals sites of interaction with 16S rRNA.

Understanding the process whereby the ribosome translates the genetic code into protein molecules will ultimately require high-resolution structural information, and we report here the first crystal structure of a protein from the small ribosomal subunit. This protein, S5, has a molecular mass of 17,500 and is highly conserved in all lifeforms. The molecule contains two distinct alpha/beta domains that have structural similarities to several other proteins that are components of ribonucleoprotein complexes. Mutations in S5 result in several phenotypes which suggest that S5 may have a role in translational fidelity and translocation. These include ribosome ambiguity or ram, reversion from streptomycin dependence and resistance to spectinomycin. Also, a cold-sensitive, spectinomycin-resistant mutant of S5 has been identified which is defective in initiation. Here we show that these mutations map to two distinct regions of the molecule which seem to be sites of interaction with ribosomal RNA. A structure/function analysis of the molecule reveals discrepancies with current models of the 30S subunit.

Amino Acid Sequence↗

PubMed1992

Elementary methods for the analysis of dichotomous outcomes in unselected samples of twins.

This paper presents an elementary statistical method for analyzing dichotomous outcomes in unselected samples of twin pairs using stratified estimators of the odds ratio. The methodology begins by first randomly designating one member of each twin pair as an "index" twin and the other member as the "co-twin." Stratifying on zygosity, odds ratios are used to measure the association between disease in the index twin and disease in the co-twin. From these zygosity-specific tables we calculate the Woolf-Haldane estimator of the common odds ratio (psi F, the weighted average of the zygosity-specific odds ratios), the Mantel-Haenszel test statistic (chi 2M-H) for the common odds ratio, and a test (chi 2G) for the difference in the zygosity-specific odds ratios. In this application, psi F provides an estimate of the familial association for disease and the accompanying chi 2M-H provides a test of the null hypothesis, psi F = 1 (i.e., there is no evidence for a familial influence on disease). The chi 2G is a test of the null hypothesis that psi MZ = psi DZ; a significant value for chi 2G suggests a genetic influence on disease (assuming that the observed odds ratios follow a pattern where psi MZ greater than psi DZ). A new test statistic (chi 2c) is proposed that incorporates the expectation that psi MZ = psi 2DZ under a purely additive genetic model with no common environmental effects. A significant value of chi c2 indicates that the different odds ratios across zygosity are partly due to common environmental influences. Conversely, a nonsignificant value of chi 2c is an indication that the zygosity-specific odds ratios are due solely to additive genetic effects and not to common environment. This basic approach is extended to examine the effects of measured indicators of the specific environment and the assessment of certain forms of gene by environment interaction. All of the methods are easily understood, highly flexible, readily computed using a hand calculator, and incorporate the inherent genetic information contained within twin samples.

Data Interpretation, Statistical↗

PubMed1992

The menstrual cycle: relations of biophysical and hormonal determinations in normal women of reproductive age.

Appropriate relationships between circulating levels of 17 beta-E2 and P are necessary for normal endometrial development and blastocyst implantation. The aim of this study was to relate biophysical and biochemical measurements obtained during the menstrual cycle of six healthy women in 25 menstrual cycles. One hundred eighteen vaginosonographic determinations of the endometrial thickness together with serum E2 and P assays were performed at 5-day intervals. The three parameters studied were standardized by the formula: observation minus mean determination for that parameter divided by the standard deviation of that parameter. Using this common unit, the physiological relations between ultrasonographically determined endometrial thickness, E2, and P were readily seen.

Adult↗

PubMed1991

Conformation of one- and two-chain high molecular weight urokinase analyzed by small-angle neutron scattering and vacuum ultraviolet circular dichroism.

The structures of one- and two-chain high molecular weight human urokinase were analyzed by small-angle neutron scattering and vacuum ultraviolet circular dichroism. Both one- and two-chain high molecular weight urokinases exhibited a radius of gyration of 31 A and a maximum dimension of 90 A. Neither parameter was affected by the presence of lysine sufficient to saturate all the lysine-binding sites in human plasminogen. These physical parameters are consistent with the sedimentation coefficient of high molecular weight urokinase and indicate that both proteins are highly asymmetric. Neither protein contained much alpha-helix or parallel beta-sheet. Most of the secondary structure was in the form of antiparallel beta-sheet and beta-turns, very similar to the secondary structure of plasminogen. The macroscopic kinetic constants, Km and kcat, for the hydrolysis of (pyroGlu-Gly-Arg-NH)2-rhodamine by two-chain high molecular weight urokinase and low molecular weight urokinase which lacks the epidermal growth factor and kringle domains were similar. These structural and kinetic data are consistent with the domains in both forms of urokinase being independent structural and functional units.

Circular Dichroism↗

PubMed1991

Conformation of Lys-plasminogen and the kringle 1-3 fragment of plasminogen analyzed by small-angle neutron scattering.

Native human Glu-plasminogen (Glu1-Asn791) was previously shown to have a radius of gyration of 39 A and a shape best described by a prolate ellipsoid [Mangel, W. F., Lin, B., & Ramakrishnan, V. (1990) Science 248, 69-73]. Upon occupation of a weak lysine-binding site, the shape reversibly changes to that best described by a Debye random coil with a radius of gyration of 56 A. Conversion from the closed to the open form is not accompanied by any change in secondary structure, hence the closed conformation is formed by interaction between domains, the five kringles and the protease domain, and this is abolished upon conversion to the open form. Here we analyzed by small-angle neutron scattering the conformations of human Lys-plasminogen (Lys78-Asn791) and the fragment K1-3 that contains the first three kringles of plasminogen (Tyr80-Val338 or Tyr80-Val354). The shape of Lys-plasminogen was best described by a Debye random coil with a radius of gyration of 51 A, and occupation of its lysine-binding sites by 6-aminohexanoic acid did not dramatically alter its conformation. Thus Lys-plasminogen was in the open form, similar to that of Glu-plasminogen with its lysine-binding sites occupied. The fragment K1-3 in the absence or presence of 6-aminohexanoic acid had a shape best described equally either by an elongated prolate ellipsoid or by a Debye random coil, with a radius of gyration of 29 A. Our model for the two forms of plasminogen is that, in the closed form, domain interaction generates a compact, almost globular, structure.(ABSTRACT TRUNCATED AT 250 WORDS)

Amino Acid Sequence↗

PubMed1991

Cloning, sequencing, and overexpression of genes for ribosomal proteins from Bacillus stearothermophilus.

Although a low resolution model for the arrangement of the proteins of the small and large ribosomal subunits is known, a detailed mechanistic understanding of the function of the ribosome awaits a high resolution structure of its components. While crystals have been obtained of several ribosomal proteins from Bacillus stearothermophilus, determination of atomic resolution structures of these proteins is impeded by the difficulty of obtaining large amounts of native proteins for crystallographic or NMR studies. We describe here the cloning and overexpression in Escherichia coli of the genes for ribosomal proteins S5, L6, L9, and L18 from B. stearothermophilus. S5 is extremely toxic to E. coli when overexpressed, and we have taken advantage of a new tightly regulated expression system to obtain high yields (more than 100 mg of pure protein/liter of culture) of this protein. The B. stearothermophilus S5 produced in E. coli crystallizes, and the crystals are identical to those obtained from the native protein. The crystals diffract to 2-A resolution.

Amino Acid Sequence↗

PubMed1991

Doppler evaluation of the fetoplacental circulation in the latent phase of preterm premature rupture of membranes.

Blood velocity waveform analysis by pulsed Doppler ultrasonography in preterm premature rupture of membranes is described. Reduced amounts of amniotic fluid did not impair determination of the resistance indices from the umbilical artery. Good results were obtained from the internal carotid artery in 82% of cases. Failure in measurements at the internal carotid artery occurred when fetal head engagement prevented good study planes. Ninety-three percent of umbilical artery determinations fell within the normal limits for our control population as did 89.2% of those at the internal carotid artery. No differences in the resistance indices could be attributed to the length of the latent period in prematurely ruptured membranes or subclinical chorioamnionitis. Therefore, if a mother or fetus has an underlying condition that affects fetoplacental circulation, the resulting abnormal blood velocity waveform should reflect this condition rather than the effect of ruptured membranes.

Blood Flow Velocity↗

PubMed1991

Iniencephaly with cyclopis (a case report).

Iniencephaly is a rare neural tube defect. We report a rare association of iniencephaly with cyclopia, probably the third such report in the literature.

Abnormalities, Severe Teratoid↗