Adipofascial flap harvest using endoscopic assistance.
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Endoscopy↗
Biomedical subjects
V Ramakrishnan
Publications and source records attributed to V Ramakrishnan.
Familial determinants of moderate and intense physical activity: a twin study.
This twin study estimates familial clustering of moderate and intense leisure-time physical activity and investigates quantitatively its genetic and environmental components. Study subjects are 3,344 male twin pairs aged 33-51 yr. Moderate activity levels were assessed with six questions about discretionary walking or stair climbing for exercise. Five questions assessed regular participation in specific, intense athletic activities (running, bicycling, swimming, racquet, and other sports). Familial aggregation is estimated by odds ratio of one twin engaging in an activity when his co-twin does. Monozygotic and dizygotic twin correlations were compared to estimate genetic and nongenetic sources of phenotypic variation. For each activity, the familial aggregation odds ratio was statistically significant with values between 2.9 to 4.6 for intense activities and between 1.4 to 1.9 for all moderate activities but one. Monozygotic twin correlations were higher than dizygotic, suggesting genes play a role in the observed phenotypic variation. For four questions, and a compromise scale of moderate activity, the difference between correlations was statistically significant (P < 0.05). In this cohort, much of the phenotypic variability for both moderate and intense activities is a result of familial effects. Genes may influence regular participation in specific intense exercises more than moderate activity, such as walking for exercise.
Adult↗
Dust-mite-allergen concentrations in asthmatics' bedrooms in the Quad Cities (Illinois, USA) after the Mississippi River floods of 1993.
This study aimed to measure allergens from Dermatophagoides farinae and D. pteronyssinus and to examine possible relationships of these mite allergens with flooding and other housing factors. A total of 313 dust samples were collected from the bedrooms of 57 asthmatics in 45 homes in the Quad Cities (Illinois, USA) and analyzed by ELISA for the presence of the D. farinae and D. pteronyssinus allergens. Twenty of these homes had some flooding in the last 12 months due to the Mississippi River floods of 1993 and/or other factors. The log-transformed least-squares means of allergens collected were 28 ng/m2 for the D. farinae allergen and 26 ng/m2 for the D. pteronyssinus allergen. D. farinae allergen levels were significantly higher in homes located in the valley, in homes during the summer months, in homes with furred or feathered pets, in homes which had not been flooded in the last year, and in homes where rugs had been steam-cleaned in the last 12 months. D. pteronyssinus allergen levels were significantly higher in homes located in the valley, in homes during April and July-September, in homes with furred or feathered pets, and in homes with no dehumidifier.
Allergens↗
CD11b mRNA expression in neutrophils isolated from peripheral blood and gingival crevicular fluid.
Adhesion molecule CD11b/CD18 expressed by neutrophils (PMNs) participates in cell migration and phagocytosis of C3bi derivatized bacteria. It is this phagocytic function that eliminates some of the known periodontal pathogens in periodontal pockets. In patients with advanced periodontitis, homotypic aggregation of crevicular fluid PMNs (CF-PMNs) may occur due to overexpression of CD11b/CD18 and this may lead to ineffective elimination of periodontal pathogens. We have previously shown that CF-PMNs isolated from the periodontal pockets overexpress CD11b compared to PB-PMNs. This study tested the hypotheses that (1) overexpression of surface CD11b correlates with expression of CD11b mRNA in CF-PMNs isolated from advanced periodontitis subjects, and (2) the intrinsic capacity of CD11b mRNA upregulation by PB-PMNs from periodontitis patients differs from that of control subjects. CF-PMNs and peripheral blood PMNs (PB-PMNs) were isolated from 13 subjects with healthy gingiva (control group) and 13 subjects with advanced periodontitis (patient group). The surface expression of CD11b was determined by flow cytometry and CD11b mRNA was determined by extraction of mRNA and reverse transcription to cDNA followed by DNA amplification using primers to detect a segment of the cDNA which encodes CD11b. The results of this study confirm that the surface expression of CD11b on CF-PMNs is significantly higher in periodontitis subjects vs control subjects (p = 0.03), whereas surface CD11b expression on PB-PMNs does not differ significantly between groups (p = 0.06). The level of surface CD11b expression on CF-PMNs did not correlate with the amount of mRNA present in CF-PMNs in either group (p = 0.056, 0.07 for control and periodontitis patients, respectively). Most (9 of 13) individuals in the patient group expressed CD11b mRNA whereas very few control subjects (2 of 11) had CD11b mRNA in their CF-PMNs. This difference between groups was statistically significant (p = 0.004). The capacity to upregulate CD11b mRNA upon stimulation with fMLP and/or GM-CSF was highly variable and there was no statistical difference between the 2 groups.
Adult↗
Characterization of Di-myo-Inositol-1,1(prm1)-Phosphate in the Hyperthermophilic Bacterium Thermotoga maritima.
Di-myo-inositol-1,1(prm1)-phosphate (DIP) is present at a significant concentration ((symbl)160 nmol/mg of protein) in the cytoplasm of the hyperthermophilic bacterium Thermotoga maritima. The concentration of DIP was independent of the pH of the growth medium or the cell growth phase but increased with increasing concentrations of NaCl in the growth medium, reaching a maximum ((symbl)450 nmol/mg of protein) at 0.4 to 0.6 M NaCl. A large-scale purification procedure for DIP which yields approximately 18 g of DIP per kg of cells (wet weight) is described. Purified DIP was stable at 90(deg)C for at least 5 h. The presence of DIP (50 mM) did not increase the stability at 90(deg)C of pure forms of the hydrogenase or pyruvate ferredoxin oxidoreductase of T. maritima, suggesting that DIP is not a general thermoprotectant.
Journal Article↗
Characterization of UDP amino sugars as major phosphocompounds in the hyperthermophilic archaeon Pyrococcus furiosus.
The archaeon Pyrococcus furiosus is a strictly anaerobic heterotroph that grows optimally at 100 degrees C by the fermentation of carbohydrates. It is known to contain high concentrations of novel intracellular solutes such as beta-mannosylglycerate and di-myo-inositol 1,1'-phosphate (DIP) (L. O. Martins and H. Santos, Appl. Environ. Microbiol. 61:3299-3303, 1995). Here, 31P nuclear magnetic resonance (NMR) spectroscopy was used to show that this organism also accumulates another type of phospho compound, as revealed by a major multiplet signal in the pyrophosphate region. The compounds were purified from cell extracts of P. furiosus by anion-exchange and gel filtration chromatographic procedures and were structurally analyzed by 1H, 13C, and 31P NMR spectroscopy. They were identified as two uridylated amino sugars, UDP N-acetylglucosamine and UDP N-acetylgalactosamine. Unambiguous characterizations and complete assignments of 1H and 13C resonances from such sugars have not been previously reported. In vitro 31P NMR spectroscopic analyses showed that, in contrast to DIP, which is maintained at a constant intracellular concentration (approximately 32 mM) throughout the growth phase of P. furiosus, the UDP amino sugars accumulated (to approximately 14 mM) only during the late log phase. The possible biochemical roles of these compounds in P. furiosus are discussed.
Archaea↗
Histone structure and the organization of the nucleosome.
Chromatin structure is now believed to be dynamic and intimately related with cellular processes such as transcription. Over the past few years, high-resolution structures for the histones have become available. These structures and their implications for nucleosome organization are reviewed here.
Amino Acid Sequence↗
Histone H1 and chromatin higher-order structure.
The linker histone H1, or its variants such as H5, have long thought to be involved in promoting the organization of chromatin into a higher-order structure, the 30 nm filament. However, the location of H1 in the filament, its role in filament formation and the structure of the 30 nm filament itself have all been controversial. This article reviews recent results that address these questions.
Chromatin↗
Treatment of multiwavelength anomalous diffraction data as a special case of multiple isomorphous replacement.
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Algorithms↗
Investigation with chitosan-oxalate oxidase-catalase conjugate for degrading oxalate from hyperoxaluric rat chyme.
Enteric hyperoxaluria manifests due to hyperabsorption of dietary oxalate, secondary to a variety of chronic gastrointestinal disorders. The potential use of chitosan immobilized oxalate oxidase-catalase conjugate to deplete the oxalate content of food materials, while they are in the digestive tract has been evaluated by treating rat stomach chyme with such an enzyme preparation. Oxalate oxidase, obtained from beet stem, was adsorbed on chitosan along with catalase and then cross linked with glutaraldehyde to stabilize the derivative. This chemical modification of oxalate oxidase brought about a shift in its optimal pH from 4.2 to 3.8 with a marginal increase in its K(m). Compared to native enzyme, the modified oxalate oxidase exhibited increased storage stability, higher thermal stability and enhanced resistance to proteolytic digestion and heavy metal inactivation. These improved properties of the immobilized oxalate oxidase possibly render it suitable for oral administration under hyperoxaluric conditions.
Animals↗
Ribosomal protein L9: a structure determination by the combined use of X-ray crystallography and NMR spectroscopy.
The structure of protein L9 from the Bacillus stearothernophilus ribosome has been determined at 2.5 A resolution by refinement against single crystal X-ray diffraction data with additional constraints provided by NMR data. This highly elongated protein consists of two domains separated by a nine-turn connecting helix. Conserved aromatic and positively charged amino acid residues on the surface of each domain are likely to be directly involved in binding 23 S ribosomal RNA. The shape of the protein, with its two widely spaced RNA-binding sites, suggests that it may serve as a "molecular strut", most likely playing a role in ribosome assembly and/or maintaining the catalytically active conformation of the ribosomal RNA. The combined use of X-ray and NMR data in the refinement procedure was essential in defining the N-terminal domain of the protein, which was relatively poorly determined by the X-ray data alone. In addition to resolving the ambiguities in defining the hydrophobic core and side-chain conformations with the N-terminal domain, this combined NMR-X-ray analysis provides the first detailed and accurate view of the N-terminal RNA-binding site. NMR data also showed that the N-terminal domain is stable in solution, indicated by amide protons that are protected from solvent exchange. The lack of definition of the N-terminal domain in the X-ray structure is therefore likely due to packing disorder within the crystal rather than structural instability. This combined NMR-X-ray analysis provides a useful model as to how X-ray and NMR data can be practically and logically combined in the determination of the structure of a single protein molecule.
Amino Acid Sequence↗
Structural evidence for specific S8-RNA and S8-protein interactions within the 30S ribosomal subunit: ribosomal protein S8 from Bacillus stearothermophilus at 1.9 A resolution.
BACKGROUND: Prokaryotic ribosomal protein S8 is an important RNA-binding protein that occupies a central position within the small ribosomal subunit. It interacts extensively with 16S rRNA and is crucial for the correct folding of the central domain of the rRNA. S8 also controls the synthesis of several ribosomal proteins by binding to mRNA. It binds specifically to very similar sites in the two RNA molecules. RESULTS: S8 is divided into two tightly associated domains and contains three regions that are proposed to interact with other ribosomal components: two potential RNA-binding sites, and a hydrophobic patch that may interact with a complementary hydrophobic region of S5. The N-terminal domain fold is found in several proteins including two that bind double-stranded DNA. CONCLUSIONS: These multiple RNA-binding sites are consistent with the role of S8 in organizing the central domain and agree with the latest models of the 16S RNA which show that the S8 location coincides with a region of complicated nucleic-acid structure. The presence in a wide variety of proteins of a region homologous to the N-terminal domain supports the idea that ribosomal proteins must represent some of the earliest protein molecules.
Amino Acid Sequence↗
Identification of two DNA-binding sites on the globular domain of histone H5.
The nature of the complexes of histones H1 and H5 and their globular domains (GH1 and GH5) with DNA suggested two DNA-binding sites which are likely to be the basis of the preference of H1 and H5 for the nucleosome, compared with free DNA. More recently the X-ray and NMR structures of GH5 and GH1, respectively, have identified two basic clusters on opposite sides of the domains as candidates for these sites. Removal of the positive charge at either location by mutagenesis impairs or abolishes the ability of GH5 to assemble cooperatively in 'tramline' complexes containing two DNA duplexes, suggesting impairment or loss of its ability to bind two DNA duplexes. The mutant forms of GH5 also fail to protect the additional 20 bp of nucleosomal DNA that are characteristically protected by H1, H5 and wild-type recombinant GH5. They still bind to H1/H5-depleted chromatin, but evidently inappropriately. These results confirm the existence of, and identify the major components of, two DNA-binding sites on the globular domain of histone H5, and they strongly suggest that both binding sites are required to position the globular domain correctly on the nucleosome.
Amino Acid Sequence↗
The proteinase-activated receptor 2 is induced by inflammatory mediators in human endothelial cells. Comparison with the thrombin receptor.
The proteinase-activated receptor 2 (PAR-2) belongs to the family of seven transmembrane region receptors, and, like the related thrombin receptor, it is activated by specific proteolytic cleavage of its extracellular amino terminus. It is not known which proteinase is the physiological activator of the PAR-2, but candidates can be found among the enzymes involved in the inflammatory cascade systems. Here, we have studied the effects of various mediators on the expression of the PAR-2 and the thrombin receptor in cultured human umbilical vein endothelial cells. Stimulation with the cytokines tumor necrosis factor alpha or interleukin-1 alpha as well as bacterial lipopolysaccharide elevated the expression of PAR-2 in a dose-dependent manner. The time course of induction after cytokine stimulation was similar to those published for the adhesion molecules intercellular adhesion molecule-1 and vascular cell adhesion molecule-1. After 20 h of stimulation, PAR-2 mRNA and protein levels were increased to 5-10-fold basal values, and, in the continued presence of tumor necrosis factor alpha, PAR-2 mRNA expression was found to remain elevated for up to 4 days. In contrast, the thrombin receptor gene was not induced by any of these inflammatory mediators. The responses to phorbol ester treatment also differed between the two genes. Thrombin receptor mRNA levels decreased steadily up to 20 h, whereas PAR-2 mRNA levels first rose to about 3-fold basal values at 4 h before decreasing again. Cell surface protein levels of both receptors were decreased after 20 h of phorbol ester stimulation. Elevating intracellular cAMP levels by treatment with forskolin resulted in decreased expression of both receptors, and inhibition of cAMP degradation appeared to blunt the cytokine-induced increase in PAR-2 expression. The induction of the PAR-2 by cytokine treatment supports the concept of PAR-2 involvement in the acute inflammatory response.
1-Methyl-3-isobutylxanthine↗
Latent hypoparathyroidism in children with conotruncal cardiac defects.
BACKGROUND: DiGeorge anomaly is characterized by hypoplasia or atresia of the thymus and parathyroid glands resulting in T cell-mediated deficiency, hypocalcemic hypoparathyroidism, and conotruncal cardiac defects. It usually is associated with deletions of chromosomal region 22q11. We hypothesized that the stimulated (secretory reserve) but not the constitutive secretion of parathyroid hormone would be reduced in normocalcemic children with conotruncal cardiac defects but no overt immune deficiency and would be related to the presence of a deletion in the DiGeorge chromosomal region of 22q11. METHODS AND RESULTS: Blood-ionized calcium and serum-intact parathyroid hormone were measured at baseline and seven more times during hypocalcemia induced during cardiopulmonary bypass in 22 patients and 10 control subjects with an atrial septal defect. Chromosomal deletions were detected by fluorescent in situ hybridization and DNA dosage analysis. There were no differences in basal calcium and parathyroid hormone levels between patients and control subjects. All had increased parathyroid hormone in response to hypocalcemia; despite lower calcium levels, parathyroid hormone levels were lower in patients. The parathyroid hormone secretory reserve in 14 of 22 patients was reduced compared with control subjects; 4 of the 14 had deletions. CONCLUSIONS: A significant number of children with conotruncal cardiac defects have normocalcemia and a normal constitutive level of parathyroid hormone but deficient parathyroid hormone secretory reserve; about 30% also have 22q11 deletions. Such children may be at risk for the later development of hypocalcemic hypoparathyroidism.
Adolescent↗
Linker histone-dependent DNA structure in linear mononucleosomes.
We have examined the binding of the linker histone H5 (LH) to mononucleosomes. Mononucleosomes reconstituted on short DNA fragments display a series of discrete bands on a gel corresponding to various nucleosome positions along the DNA. When a series of engineered H5s with differing extents of the C-terminal tail are bound to these mononucleosomes, the electrophoretic mobilities of the resulting complexes are altered. Not only is there a general increase in mobility upon complex formation, but there is a reduction in the differences in mobility of the most distal nucleosomes. The complexes were also visualized by electronmicroscopy. From these two complementary studies, we conclude the following. (1) Entering and exiting DNAs are uncrossed in the LH-free particles, despite a DNA wrapping of 1.65 to 1.7 turns around the histone core. This results from a bending of the entering and exiting DNA away from each other and the histone surface, presumably as a consequence of electrostatic repulsion. This confirms and extends conclusions derived from our recent examination of the same particles in 3D through cryo-electron microscopy. (2) Binding of the globular domain of H5 increases DNA wrapping to 1.8 to 1.9 turns, but fails to induce a crossing due to an accentuation of the bends. (3) The C-terminal tail of H5 bridges entering and exiting DNAs together into a four-stranded stem over a distance of about 30 bp. The occurrence of such a stem may introduce constraints on models of the 30 nm chromatin fiber.
Animals↗
Solution structure of prokaryotic ribosomal protein S17 by high-resolution NMR spectroscopy.
The solution of a primary 16S rRNA-binding ribosomal protein, S17, was investigated by two- and three-dimensional homonuclear and heteronuclear magnetic resonance spectroscopy. Almost complete chemical shift assignments for the 1H, 15N, and 13C resonances have been obtained. The NMR data have been rigorously analyzed using a combination of distance geometry, back-calculation, and simulated annealing refinement techniques, and a high-resolution three-dimensional structure has been deduced. The protein consists of a single twisted antiparallel beta-pleated sheet with Greek-key topology. The five beta-strands are connected by extended loops that are flexible compared to the beta-sheet core structure and appear not to adopt one definite conformation in solution. Two of these loops contain many of the residues that have been implicated in binding ribosomal RNA. The location and distribution of these residues and other positively charged side chains on the protein surface suggest an interaction with two distinct regions of ribosomal RNA.
Amino Acid Sequence↗
The crystal structure of ribosomal protein L14 reveals an important organizational component of the translational apparatus.
BACKGROUND: Detailed structural information on ribosomal proteins has increased our understanding of the structure, function and evolution of the ribosome. L14 is one of the most conserved ribosomal proteins and appears to have a central role in the ribonucleoprotein complex. Studies have indicated that L14 occupies a central location between the peptidyl transferase and GTPase regions of the large ribosomal subunit. RESULTS: The crystal structure of L14 from Bacillus stearothermophilus has been solved using a combination of isomorphous replacement and multiwavelength anomalous dispersion (MAD) methods. The structure comprises a five-stranded beta-barrel, a C-terminal loop region that contains two small alpha-helices, and a beta-ribbon that projects from the beta-barrel. An analysis of the structure and the conserved amino acids reveals three surface patches that probably mediate L14-RNA and L14-protein interactions within the ribosome. CONCLUSIONS: The accepted role of ribosomal proteins is to promote the folding and stabilization of ribosomal RNA. The L14 structure is consistent with this notion, and it suggests that the RNA binds in two sites. One RNA-binding site appears to recognize a distinct region of ribosomal RNA during particle assembly. The second site is smaller and may become occupied during the later compaction of the RNA. The surface hydrophobic patch is a likely site of protein-protein interaction, possibly with L19.
Amino Acid Sequence↗