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Biomedical subjects

E Arnold

Publications and source records attributed to E Arnold.

At least 19 recordsLinked to original sources
PubMed1992

Structure of HIV-1 reverse transcriptase/DNA complex at 7 A resolution showing active site locations.

AIDS, caused by human immunodeficiency virus (HIV), is one of the world's most serious health problems, with current protocols being inadequate for either prevention or successful long-term treatment. In retroviruses such as HIV, the enzyme reverse transcriptase copies the single-stranded RNA genome into double-stranded DNA that is then integrated into the chromosomes of infected cells. Reverse transcriptase is the target of the most widely used treatments for AIDS, 3'-azido-3'-deoxythymidine (AZT) and 2',3'-dideoxyinosine (ddI), but resistant strains of HIV-1 arise in patients after a relatively short time. There are several nonnucleoside inhibitors of HIV-1 reverse transcriptase, but resistance to such agents also develops rapidly. We report here the structure at 7 A resolution of a ternary complex of the HIV-1 reverse transcriptase heterodimer, a monoclonal antibody Fab fragment, and a duplex DNA template-primer. The double-stranded DNA binds in a groove on the surface of the enzyme. The electron density near one end of the DNA matches well with the known structure of the HIV-1 reverse transcriptase RNase H domain. At the opposite end of the DNA, a mercurated derivative of UTP has been localized by difference Fourier methods, allowing tentative identification of the polymerase nucleoside triphosphate binding site. We also determined the structure of the reverse transcriptase/Fab complex in the absence of template-primer to compare the bound and free forms of the enzyme. The presence of DNA correlates with movement of protein electron density in the vicinity of the putative template-primer binding groove. These results have important implications for developing improved inhibitors of reverse transcriptase for the treatment of AIDS.

Base Sequence↗
PubMed1992

Crystallization and preliminary X-ray diffraction studies of coxsackievirus B1.

Preparations of coxsackievirus B1 (CVB1) derived from an infectious cDNA clone have been crystallized in multiple crystal forms. Using high intensity synchrotron radiation, an orthorhombic form of the crystals was shown to diffract X-rays to at least 2.9 A resolution. The unit cell has a primitive lattice with dimensions a = 323 A, b = 450 A, and c = 522 A. A crystallographic asymmetric unit of these CVB1 crystals probably contains an entire virus particle, implying the presence of 60-fold non-crystallographic redundancy. This CVB1 crystal form appears to be suitable for high-resolution structure determination by X-ray crystallography.

Crystallography↗
PubMed1992

Is the tyramine test for depressive illness useful in elderly patients?

There were no significant differences in tyramine sulphate excretion following tyramine ingestion between elderly depressed, demented or control patient groups, in contrast with younger subjects where this test is a trait marker for unipolar endogenous depression. There are inherent problems in urine collection studies in the elderly and the results may have been influenced by the medication that elderly patients have to take for other disorders. This study suggests that the tyramine test is unlikely to be of clinical usefulness in the over 65 age group.

Aged↗
PubMed1991

Crystals of a ternary complex of human immunodeficiency virus type 1 reverse transcriptase with a monoclonal antibody Fab fragment and double-stranded DNA diffract x-rays to 3.5-A resolution.

Two crystal forms of complexes have been grown that contain human immunodeficiency virus type 1 reverse transcriptase and a monoclonal antibody Fab fragment. One of the crystal forms (form II, space group P3112, a = 168.7 A, c = 220.3 A) diffracts x-rays to 3.5-A resolution and appears suitable for moderate-resolution structure determination. The form II crystals have the unusual property that their maximum resolution of diffraction and resistance to radiation damage are enhanced by either crystallization in the presence of or soaking with double-stranded DNA primer-template mimics. These crystals may permit structural studies of catalytically relevant complexes and eventually enable us to experimentally observe successive steps in the reverse transcription process.

Antibodies, Monoclonal↗
PubMed1991

Crystallization and preliminary X-ray diffraction analysis of oucleoside diphosphate kinase from Myxococcus xanthus.

Nucleoside diphosphate (NDP) kinase catalyzes the transfer of the gamma-phosphate from a nucleoside triphosphate to a nucleoside diphosphate. Human and rodent forms of this enzyme have been shown to be suppressors of metastasis. Crystals that diffract X-rays to high resolution have been obtained for the recombinant Myxococcus xanthus NDP kinase expressed in and purified from Escherichia coli. Two crystal forms have been obtained. Both forms are orthorhombic, space group I222 (or I2(1)2(1)2(1)) with a = 267.1 A, b = 74.0 A and c = 75.1 A for form I and a = 53.5 A, b = 74.0 A and c = 75.1 A for form II. Form I appears to have five molecules in the asymmetric unit approximately related to each other by a translation of 0.2 along the a axis. Diffraction data have been recorded to 1.9 A for form I and to 2.2 A for form II.

Crystallization↗
PubMed1991

HIV reverse transcriptase structure-function relationships.

HIV reverse transcriptase (RT) is the target of the most widely used treatments for AIDS. Biochemical and mutagenesis studies performed on HIV-1 RT are reviewed in light of the enzyme's structure and functions. Features described include domain arrangement, dimerization, proteolytic processing, and specific recognition of the priming tRNA. Possible regions of functional importance as determined by comparative amino acid sequence analysis and by site-directed mutagenesis are identified. Among the conclusions of the analysis is the unexpected realization that the substrate for proteolytic maturation of the HIV-1 RT p66/p66 homodimer to the p66/p51 heterodimer is most likely an unfolded RNase H domain. In addition, the current progress in crystallization and structure determination of HIV-1 RT is described. Finally, a functional-model of the active reverse transcription complex is presented.

Amino Acid Sequence↗
PubMed1990

Analysis of the structure of a common cold virus, human rhinovirus 14, refined at a resolution of 3.0 A.

Human rhinovirus 14 has a pseudo T = 3 icosahedral structure in which 60 copies of the three larger capsid proteins VP1, VP2 and VP3 are arranged in an icosahedral surface lattice, reminiscent of T = 3 viruses such as tomato bushy stunt virus and southern bean mosaic virus. The overall secondary and tertiary structures of VP1, VP2 and VP3 are very similar. The structure of human rhinovirus 14, which was refined at a resolution of 3.0 A [R = 0.16 for reflections with F greater than 3 sigma(F)], is here analyzed in detail. Quantitative analysis of the surface areas of contact (proportional to hydrophobic free energy of association) supports the previously assigned arrangement within the promoter, in which interactions between VP1 and VP3 predominate. Major contacts among VP1, VP2 and VP3 are between the beta-barrel moieties. VP4 is associated with the capsid interior by a distributed network of contacts with VP1, VP2 and VP3 within a promoter. As the virion assembly proceeds, the solvent-accessible surface area becomes increasingly hydrophilic in character. A mixed parallel and antiparallel seven-stranded sheet is composed of the beta C, beta H, beta E and beta F strands of VP3 in one pentamer and beta A1 and beta A2 of VP2 and the VP1 amino terminus in another pentamer. This association plays an essential role in holding pentamers together in the mature virion as this contact region includes more than half of the total short non-bonded contacts between pentamers. Contacts between protomers within pentamers are more extensive than the contacts between pentamers, accounting in part for the stability of pentamers. The previously identified immunogenic regions are correlated with high solvent accessibility, accessibility to large probes and also high thermal parameters. Surface residues in the canyon, the putative cellular receptor recognition site, have lower thermal parameters than other portions of the human rhinovirus 14 surface. Many of the water molecules in the ordered solvent model are located at subunit interfaces. A number of unusual crevices exist in the protein shell of human rhinovirus 14, including the hydrophobic pocket in VP1 which is the locus of binding for the WIN antiviral agents. These may be required for conformational flexibility during assembly and disassembly. The structures of the beta-barrels of human rhinovirus 14 VP1, VP2 and VP3 are compared with each other and with the southern bean mosaic virus coat protein.

Amino Acid Sequence↗
PubMed1989

[The viewpoint of the elderly on the problems of aging].

Members of an university for the elderly conducted a survey among 730 elderly people in order to ascertain their living and housing conditions, their familiar and social relationship, their state of health, and how they envisaged their future should their independence become limited. This article describes the methodology of the study, the rich human and social experience gained by the researchers and it indicates the characteristics of the people surveyed, a high proportion of whom were very old and living alone and would thus be in a particularly precarious situation if their independence became limited owing to age or illness.

Activities of Daily Living↗
PubMed1989

Diethylthiocarbamic acid methyl ester. A potent inhibitor of aldehyde dehydrogenase found in rats treated with disulfiram or diethyldithiocarbamic acid methyl ester.

Rats were treated with disulfiram (Antabuse, DSF) or its metabolite diethyldithiocarbamic acid methyl ester (Me-DDC) and challenged with ethanol. The blood pressure response to ethanol was followed and blood was analyzed for DSF, Me-DDC and diethyldithiocarbamic acid (DDC). The rat liver aldehyde dehydrogenase (ALDH) isozyme activities were measured 2 hr after the ethanol challenge. Both treatments produced a significant fall in the blood pressure when challenged with ethanol, probably caused by a marked decrease in hepatocyte low Km and high Km activities. The mean plasma concentration ranges of Me-DDC and DDC were found to be 49-1241 nmol/l and 182-841 nmol/l, respectively, whereas DSF was undetectable. In addition, it was found that inactivation of hepatocyte low Km ALDH activity was dependent on preoxidation of Me-DDC by the microsomal cytochrome P-450 mixed function oxidases. Me-DDC was found to be oxidized under aerobic conditions in the presence of NADP to form diethylthiocarbamic acid methyl ester (Me-DTC). The structure was confirmed from its MS/EI fragmentation spectrum. Me-DTC was found to be a potent inhibitor of low Km ALDH when added to rat liver homogenate. The compound was also identified as a metabolite in rat blood collected from the DSF and Me-DDC treated rats, and in blood from human alcoholics on DSF treatment. Me-DTC appears to be more selective for the low Km isozymes whereas the opposite seems to be the case for the hydrolytic product, DTC.

Aldehyde Dehydrogenase↗
PubMed1989

[Descriptive single case analyses--a strategy for studying effectiveness correlations in psychotherapy].

A method of data analysis based on single case studies is investigated. It is intended to lead to empirically grounded hypotheses about the differences in effects of various therapeutic methods. The data bases are 16 documented therapies, eight of which are client-centered therapies and interactional behavior therapies respectively. Descriptive single case analyses were carried out in four steps: 1. documentation of single cases; 2. description of individual patterns of therapeutic effects; 3. comparisons within treatment groups; 4. comparisons between treatment groups. Results show that individual differences within groups exceed similarities, so that there is little evidence of a strong influence of the treatment method on individual patterns of therapeutic effects. This result leads to the conclusion that a typology of therapeutic processes should be based on more factors than the treatment method alone.

Behavior Therapy↗
PubMed1988

The use of molecular-replacement phases for the refinement of the human rhinovirus 14 structure.

The structure of human rhinovirus 14 has been refined, by the method of restrained least squares, to an R factor of 0.16 for various random samples between 6 and 3 A resolution with F greater than 3 sigma (F). As a first step the non-crystallographic symmetry parameters were optimized using the initial atomic model in a rigid-body refinement procedure. Phase determination by the molecular-replacement phase extension and refinement procedure was continued to 2.94 A resolution, employing the improved non-crystallographic symmetry operators. The resultant structure-factor phases and weights, together with the measured amplitudes, constituted the X-ray observations used in the restrained refinement. The Hendrickson-Konnert program system [Konnert & Hendrickson (1980). Acta Cryst. A36, 344-350] was modified to incorporate non-crystallographic symmetry constrains and structure-factor phases as observations. The non-bonded contacts between subunits related by non-crystallographic symmetry were also restrained.

Chemical Phenomena↗
PubMed1988

Ischemia-induced alterations of mitochondrial structure and function in brain, liver, and heart muscle of young and senescent rats.

Young and senescent rats (3 and 28-30 months old) were subjected to complete ischemia at 37 degrees C in order to study function and structure of mitochondria isolated from liver, heart muscle, and brain. The rates of energy-coupled respiration and ATP synthesis were found to decrease progressively in relation to time of ischemia. The respiratory rates in the absence of ADP (state 4 respiration) did not increase after exposure to ischemia, suggesting that ischemia primarily affects electron transport rather than the energy coupling system. Mitochondria of heart muscle were more affected by ischemia than mitochondria of brain and liver. Liver and heart muscle mitochondria obtained from young rats were found to be slightly more sensitive to short periods of ischemia than those isolated from senescent animals.

Adenosine Triphosphate↗
PubMed1987

The atomic structure of Mengo virus at 3.0 A resolution.

The structure of Mengo virus, a representative member of the cardio picornaviruses, is substantially different from the structures of rhino- and polioviruses. The structure of Mengo virus was solved with the use of human rhinovirus 14 as an 8 A resolution structural approximation. Phase information was then extended to 3 A resolution by use of the icosahedral symmetry. This procedure gives promise that many other virus structures also can be determined without the use of the isomorphous replacement technique. Although the organization of the major capsid proteins VP1, VP2, and VP3 of Mengo virus is essentially the same as in rhino- and polioviruses, large insertions and deletions, mostly in VP1, radically alter the surface features. In particular, the putative receptor binding "canyon" of human rhinovirus 14 becomes a deep "pit" in Mengo virus because of polypeptide insertions in VP1 that fill part of the canyon. The minor capsid peptide, VP4, is completely internal in Mengo virus, but its association with the other capsid proteins is substantially different from that in rhino- or poliovirus. However, its carboxyl terminus is located at a position similar to that in human rhinovirus 14 and poliovirus, suggesting the same autocatalytic cleavage of VP0 to VP4 and VP2 takes place during assembly in all these picornaviruses.

Antigens, Viral↗
PubMed1987

Comparison of the three-dimensional structure of two human rhinoviruses (HRV2 and HRV14).

An attempt has been made to build a model of human rhinovirus 2 (HRV2) based on the known human rhinovirus 14 (HRV14) structure. HRV2 was selected because its amino acid sequence is known and because it belongs to the minor rhinovirus receptor class as compared to HRV14, which belongs to the major class. Initial alignment of HRV2 with HRV14 based on the primary sequence and the knowledge of the three-dimensional structure of HRV14 showed that the most probable position of the majority of insertions and deletions occurred in the vicinity of the neutralizing immunogenic sites (NIm). Out of a total of 855 amino acids present in one copy of each of the capsid proteins VP1 through VP4 of HRV14, 411 are different between the two viruses. There are also 6 amino acid residues inserted and 14 residues deleted in HRV2 relative to HRV14. Examination of amino acid interactions showed several cases of conservation of function, e.g., salt bridges or the filling of restricted space. The largest variation amongst the residues lining the canyon, the putative receptor binding site, was in the carboxy-terminal residues of VP1.

Amino Acid Sequence↗