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Biomedical subjects

R C Thompson

Publications and source records attributed to R C Thompson.

At least 325 records · Page 18Linked to original sources
PubMed1986

Isolation and sequence of a human gene encoding a potent inhibitor of leukocyte proteases.

We report the isolation of the human gene encoding an inhibitor of neutrophil elastase and cathepsin G. We have sequenced the gene and a cDNA clone isolated from human parotid tissue. The protein encoded by this gene appears to contain two functional domains, one having a trypsin inhibitory site and the other an elastase inhibitory site. The two-domain structure of the protein is reflected in the organization of the gene, with each domain represented by a separate exon. We have also noted that the intervening sequence separating the trypsin-inhibitor-exon and the elastase-inhibitor-exon is flanked by eleven base-pair direct repeats, suggesting that this intron may have been generated by a transposition-type event.

Base Sequence↗
PubMed1986

Effect of replacing uridine 33 in yeast tRNAPhe on the reaction with ribosomes.

We have determined several kinetic parameters for the reaction of poly(U)-programmed ribosomes with ternary complexes of elongation factor Tu, GTP, and yeast Phe-tRNA analogs with different bases substituted for uridine in position 33. These analogs test whether disruption of the hydrogen bonds normally formed by uridine 33 and steric crowding in the anticodon loop are detrimental to tRNA function on the ribosome. Single-turnover kinetic studies of the reaction of these ternary complexes with ribosomes show that these Phe-tRNA analogs decrease the apparent rate of GTP hydrolysis (kGTP) and the ratio of peptide formed to GTP hydrolyzed. Thus, the substitution of uridine 33 affects not only the selection of a ternary complex by the ribosome but also the selection of an aminoacyl-tRNA in the proofreading reaction. The effects become greater as first one, and then the other, H-bond is disrupted. Steric crowding in the anticodon loop is also important, but does not have as great an effect on the rate constants. An analysis of the elementary rate constants which comprise the rate constant, kGTP, demonstrates that the reduction in kGTP results from a decreased rate of ternary complex association with the ribosome (k1) and that there is little or no effect on the rate of GTP cleavage (k2). An analysis of the rate constants involved in proofreading shows that all the modified (tRNAs have increased rates of aminoacyl-tRNA rejection (k4) but that the rate of peptide bond formation (k3) is unaffected.

Base Sequence↗
PubMed1986

Magnetic resonance imaging during acute myocardial infarction.

Experimental canine studies have demonstrated the potential of magnetic resonance imaging (MRI) for detecting and characterizing acute myocardial infarction (AMI) in humans. Accordingly, electrocardiographic-gated spin-echo MR images of the left ventricular short axis were obtained in 34 patients a mean of 11 +/- 6 days (range 3 to 30) after AMI. This imaging technique allowed division of the left ventricle into segments corresponding to the left ventricular segments on angiography. Patients were separated into 2 groups; the first 16 patients (group I) were examined using a variety of imaging techniques. Information derived from this experience resulted in a standard imaging protocol and development of criteria for the presence of AMI. The imaging protocol and interpretation criteria were used in the assessment of a subsequent group of 18 patients (group II). Of the 14 patients in group II with satisfactory image quality, all showed an increase in myocardial signal intensity consistent with an AMI. In addition, the anterior or inferior location of the abnormal MR segments corresponded to the electrocardiographic infarct location. MR segments showing increased signal intensity corresponded with severely hypokinetic or akinetic segments on the left ventriculogram in 8 patients having both procedures. In a group of volunteers who underwent imaging and whose images were interpreted in the same manner as those of the patients with AMI, 1 of 9 subjects had regional variation in myocardial signal intensity compatible with an AMI. In summary, AMI is readily detected, located and characterized by electrocardiographic-gated MRI. These findings suggest that MRI techniques may have a role in the evaluation of AMI in humans.

Adult↗
PubMed1986

The reaction of ribosomes with elongation factor Tu.GTP complexes. Aminoacyl-tRNA-independent reactions in the elongation cycle determine the accuracy of protein synthesis.

The fidelity of protein synthesis depends on the rate constants for the reaction of ribosomes with ternary complexes of elongation factor Tu (EF-Tu), GTP, and aminoacyl (aa)-tRNA. By measuring the rate constants for the reaction of poly(U)-programmed ribosomes with a binary complex of elongation factor (EF-Tu) and GTP we have shown that two of the key rate constants in the former reaction are determined exclusively by ribosome-EF-Tu interactions and are not affected by the aa-tRNA. These are the rate constant for GTP hydrolysis, which plays an important role in the fidelity of ternary complex selection by the ribosome, and the rate constant for EF-Tu.GDP dissociation from the ribosome, which plays an equally important role in subsequent proofreading of the aa-tRNA. We conclude that the fidelities of ternary complex selection and proofreading are fundamentally dependent on ribosome-EF-Tu interactions. These interactions determine the absolute value of the rate constants for GTP hydrolysis and EF-Tu.GDP dissociation. The ribosome then uses these rate constants as internal standards to measure, respectively, the rate constants for ternary complex and aa-tRNA dissociation from the ribosome. These rates, in turn, are highly dependent on whether the ternary complex and aa-tRNA are cognate or near-cognate to the codon being translated.

Escherichia coli↗
PubMed1986

Guanosine 5'-O-(3-thiotriphosphate) as an analog of GTP in protein biosynthesis. The effects of temperature and polycations on the accuracy of initial recognition of aminoacyl-tRNA ternary complexes by ribosomes.

Guanosine 5'-O-(3-thio)triphosphate (GTP gamma S) is a good analog of GTP in the reactions leading to the formation of a peptide bond in protein biosynthesis. It forms binary and ternary complexes with elongation factor Tu (EF-Tu), and with EF-Tu and aminoacyl-tRNA (aa-tRNA). In addition, it stimulates aa-tRNA binding to ribosomes. Although GTP gamma S hydrolysis is more than three orders of magnitude slower than GTP hydrolysis, both reactions are dependent on the formation of a noncovalent complex (RS X TC) between mRNA-programmed ribosomes and ternary complex, and the complexes resulting from that hydrolysis are intermediates in peptide formation. The rate of dissociation of the ribosome X EF-Tu X GTP gamma S X aa-tRNA complex was determined from the rate of labeled peptide formation in the presence of an unlabeled ternary complex chase. This rate (2.2 X 10(-3) s-1) is similar to that determined previously (Thompson, R.C., and Karim, A.M. (1982) Proc. Natl. Acad. Sci. U.S.A. 79, 4922-4926) from the progress of GTP gamma S hydrolysis. The effects of temperature and polycation concentration on this rate constant and that for GTP gamma S hydrolysis are reported. The rate constants measured are consistent with a kinetic rather than thermodynamic limit on the accuracy of the aa-tRNA selection in vivo.

Escherichia coli↗
PubMed1986

Praziquantel adversely affects protoscoleces of Echinococcus granulosus in vitro.

Praziquantel was shown to have adverse effects on protoscoleces of Echinococcus granulosus in vitro. Exposure to one dose of praziquantel at a concentration of 10 micrograms/ml caused protoscoleces to die within 12 to 15 days. Protoscolecidal effects were more marked when cultures were exposed to the drug continuously by the addition of multiple doses. On the basis of these observations, there is a need to reappraise the metacestocidal potential of praziquantel, particularly with regard to the development of new drug regimes employing multiple dose or sustained release schedules. Possible reasons for the reduced susceptibility of protoscoleces and juvenile worms to praziquantel observed in this study are discussed.

Animals↗
PubMed1986

Isolation, properties, and complete amino acid sequence of human secretory leukocyte protease inhibitor, a potent inhibitor of leukocyte elastase.

A potent inhibitor of human leukocyte elastase (EC 3.4.21.37) and cathepsin G (EC 3.4.21.20) and of human trypsin (EC 3.4.21.4) has been purified from human parotid secretions. The complete amino acid sequence of this protein has been determined. The sequence suggests that the protein has two domains of about 54 amino acids, each of which contains four disulfide bonds. On the basis of a limited homology to other protease inhibitors, the antielastase and antitrypsin activities are thought to be properties of the C-terminal and N-terminal domains, respectively. The affinity of the inhibitor for leukocyte elastase is very high, suggesting a functional role for the protein in preventing elastase-mediated damage to oral and possibly other mucosal tissues.

Amino Acid Sequence↗
PubMed1986

Elongation factor Tu.guanosine 3'-diphosphate 5'-diphosphate complex increases the fidelity of proofreading in protein biosynthesis: mechanism for reducing translational errors introduced by amino acid starvation.

Complexes of elongation factor Tu (EF-Tu) with guanosine 3'-diphosphate 5'-diphosphate (ppGpp) bind to ribosomes where they slow the incorporation of aminoacyl-tRNAs into protein by inhibiting both the binding of aminoacyl-tRNA.EF-Tu.GTP ternary complexes and the formation of peptide bonds. The latter action increases the time available for aminoacyl-tRNA rejection by the ribosome and, therefore, increases the effectiveness of proofreading. Synthesis of ppGpp and the formation of EF-Tu.ppGpp occur in vivo in response to amino acid starvation. Our finding, therefore, suggests an explanation for the otherwise puzzling observation that amino acid starvation has, at most, a moderate effect on the fidelity of protein synthesis in wild-type Escherichia coli. We suggest that an EF-Tu.ppGpp-induced increase in the effectiveness of proofreading buffers the overall translational fidelity of these cells against amino acid starvation-induced errors in initial selection of aminoacyl-tRNA ternary complexes.

Escherichia coli↗
PubMed1986

Primary structure and cDNA cloning of human fibroblast collagenase inhibitor.

We report the primary structure and cDNA cloning of human fibroblast collagenase inhibitor, a glycoprotein that appears to play a central role in modulating the activity of a number of metalloendoproteases of connective tissue origin including collagenase, gelatinase, and proteoglycanase. Secreted human fibroblast collagenase inhibitor was purified and subjected to automated Edman degradation. The secreted protein consists of 184 amino acid residues; it contains two sites of N-linked oligosaccharide linkage and six disulfide bonds. Synthetic oligonucleotide probes based on selected amino acid sequences of the inhibitor were used to screen a lambda gt10 cDNA library from a human fibroblast line. Two overlapping cDNA clones were characterized to determine the complete coding and noncoding sequences of the specific mRNA. The amino acid sequence deduced from the nucleotide sequence agrees with that determined by protein sequencing. One clone appears to contain the complete 5' end and, in addition, the cDNA sequence predicts a 23-amino acid leader peptide. The other clone represents the 3' end of the mature message and includes a short poly(A)+ tract. This 3' sequence is remarkably similar to a reported cDNA encoding part of the protein derived from mouse fibroblast poly(A)+ RNA. However, this inhibitor has no substantial homology with previously sequenced protease inhibitors.

Amino Acid Sequence↗
PubMed1986

Synaptosomal transport of radiolabel from N-acetyl-aspartyl-[3H]glutamate suggests a mechanism of inactivation of an excitatory neuropeptide.

This study was undertaken to explore in synaptosomal preparations the disposition of N-acetyl-aspartyl-glutamate (NAAG), an endogenous acidic dipeptide neurotransmitter candidate. Radiolabel from N-acetyl-aspartyl[3H]glutamate was taken up rapidly into an osmotically sensitive compartment by rat brain synaptosomal preparations in a sodium-, temperature-, and time-dependent manner. HPLC analysis of the accumulated radiolabel indicated that the bulk of the tritium cochromatographed with glutamic acid and not with NAAG. In contrast, [14C]NAAG, labeled on the N-terminal acetate, was not taken up by the synaptosomal preparation. All effective inhibitors of synaptosomal, Na+-dependent [3H]glutamate uptake were found to exhibit similar potency in inhibiting uptake of tritium derived from [3H]NAAG. However, certain alpha-linked acidic dipeptides, structurally similar to NAAG, as well as the potent convulsant quisqualic acid inhibited synaptosomal transport of [3H]NAAG but were ineffective as inhibitors of [3H]glutamate transport. Together with a demonstration of disparities between the regional accumulation of radiolabel from [3H]NAAG and high-affinity [3H]glutamate uptake, these data suggest the presence in brain of a specific peptidase targeting carboxy-terminal glutamate-containing dipeptides that may be coupled to the Na+-dependent glutamate transporter. These findings provide a possible mechanism for NAAG inactivation subsequent to its release from nerve endings.

Amino Acids↗
PubMed1986

The prevalence of Giardia in dogs and cats in Perth, Western Australia.

A survey of dogs and cats in the Perth metropolitan area revealed a high prevalence of Giardia. Overall, 21% of 333 dogs and 14% of 226 cats were infected. More dogs and cats from refuges and breeding establishments were infected than household pets, although among the latter a significant number of dogs (9%) and cats (8%) was infected. Giardia did not show any breed or sex predisposition but prevalence was higher in young animals. The species of Giardia present in dogs and cats was identified as G. duodenalis, which is the same as that affecting man. The potential significance of this animal reservoir of infection to man is discussed in the light of increasing evidence that Giardia is a zoonosis.

Animals↗
PubMed1986

Osteosarcoma: role of CT in limb-sparing treatment.

The computed tomographic (CT) examinations of 17 patients undergoing evaluation for limb-sparing treatment of osteosarcoma were evaluated. In all cases information from CT directly affected the operative approach. The CT findings that helped in planning the operative approach included determination of intramedullary extent and evidence of "skip" metastases, direct extension into an adjacent joint, and neurovascular compromise. Performing CT of the affected bone at 1-cm intervals from the joint above through the joint below and including the contralateral side in the field of view allow precise localization of findings that can direct the operative approach. Close and contiguous scanning intervals allow identification of small intramedullary "skip" metastases that affect the resection margins.

Adolescent↗
PubMed1985

The rate of cleavage of GTP on the binding of Phe-tRNA.elongation factor Tu.GTP to poly(U)-programmed ribosomes of Escherichia coli.

Interaction of Phe-tRNA.elongation factor Tu.GTP with poly(U)-programmed ribosomes containing an occupied P site can be described by a three-step kinetic mechanism. Initial binding is followed by the cleavage of GTP, and then a new peptide bond is formed. Rate constants controlling the first and third of these reactions are known, but only a lower limit for the rate constant of the cleavage step has been reported. We have determined this rate constant to be 20 s-1 at 5 degrees C, 30 s-1 at 15 degrees C, and 50 s-1 at 25 degrees C. This is much faster than the reverse step of the initial binding process and implies that the intrinsic accuracy of the ribosome in the initial selection step is sacrificed in favor of speed. The similarity of the kinetic and chemical mechanism of this GTP cleavage step with other nucleoside 5'-triphosphatases is discussed.

Escherichia coli↗