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Biomedical subjects

L Chapman

Publications and source records attributed to L Chapman.

At least 19 recordsLinked to original sources
PubMed1996

Towards an understanding of protein-DNA recognition.

Understanding how proteins recognize DNA in a sequence-specific manner is central to our understanding of the regulation of transcription and other cellular processes. In this article we review the principles of DNA recognition that have emerged from the large number of high-resolution crystal structures determined over the last 10 years. The DNA-binding domains of transcription factors exhibit surprisingly diverse protein architectures, yet all achieve a precise complementarity of shape facilitating specific chemical recognition of their particular DNA targets. Although general rules for recognition can be derived, the complex nature of the recognition mechanism precludes a simple recognition code. In particular, it has become evident that the structure and flexibility of DNA and contacts mediated by water molecules contribute to the recognition process. Nevertheless, based on known structures it has proven possible to design proteins with novel recognition specificities. Despite this considerable practical success, the thermodynamic and kinetic properties of protein/DNA recognition remain poorly understood.

Amino Acid Sequence↗
PubMed1996

The crystal structure of the DNA-binding domain of yeast RAP1 in complex with telomeric DNA.

Telomeres, the nucleoprotein complexes at the ends of eukaryotic chromosomes, are essential for chromosome stability. In the yeast S. cerevisiae, telomeric DNA is bound in a sequence-specific manner by RAP1, a multifunctional protein also involved in transcriptional regulation. Here we report the crystal structure of the DNA-binding domain of RAP1 in complex with telomeric DNA site at 2.25 A resolution. The protein contains two similar domains that bind DNA in a tandem orientation, recognizing a tandemly repeated DNA sequence. The domains are structurally related to the homeodomain and the proto-oncogene Myb, but show novel features in their DNA-binding mode. A structured linker between the domains and a long C-terminal tail contribute to the binding specificity. This structure provides insight into the recognition of the conserved telomeric DNA sequences by a protein.

Amino Acid Sequence↗
PubMed1996

Hantavirus infections in the United States: diagnosis and treatment.

Definitive diagnosis of acute hantavirus infection requires demonstration of specific IgG and IgM antibodies and a compatible clinical course; most persons with acute illness will also have viral RNA in PBMC which can be detected after RT-PCR amplification. Characteristic hematologic and hemodynamic findings are helpful in establishing a presumptive diagnosis of HPS in persons with cardiopulmonary symptoms and signs while awaiting the results of serologic testing and PCR. Diagnosis during the prodromal phase is difficult although very limited data suggest that most these patients also have hantavirus IgG and IgM antibodies. Patients with suspected hantavirus infection should be treated in a critical care unit, as sudden hemodynamic deterioration can occur at any time. Hemodynamic monitoring is encouraged in patients with cardiopulmonary symptoms and signs, and inotropic support and vasopressors should be used rather than fluids. An uncontrolled study of ribavirin therapy was inconclusive and a placebo-controlled trial is therefore planned. The latter study aims to enroll persons with prodromal findings and early cardiopulmonary disease.

Animals↗
PubMed1996

Cloning of cDNAs from a mammalian expression library by a direct selection-amplification method.

A sensitive method was devised for cloning cDNAs from a mammalian expression library based on single-cell detection and selection of transfected cells. The method is applicable for cloning cell-surface or cytoplasmic proteins for which a detection assay, such as immunofluorescence or immunohistochemical reactivity, exists. The widely used eukaryotic expression vector cdm8 is utilized, and the method is demonstrated using the gene for C-CAM, a liver glycoprotein adhesion molecule. After transfection, cells are plated out on a collagen gel substrate to allow retrieval of selected cells. Simultaneous fixation and permeabilization with acetone permits immunological and histochemical detection of cell-surface and cytoplasmic proteins without loss of plasmid vector. Inserts of interest are recovered by PCR with vector primers.

Adenosine Triphosphatases↗
PubMed1995

The oestrogen receptor recognizes an imperfectly palindromic response element through an alternative side-chain conformation.

BACKGROUND: Structural studies of protein-DNA complexes have tended to give the impression that DNA recognition requires a unique molecular interface. However, many proteins recognize DNA targets that differ from what is thought to be their ideal target sequence. The steroid hormone receptors illustrate this problem in recognition rather well, since consensus DNA targets are rare. RESULTS: Here we describe the structure, at 2.6 A resolution, of a complex between a dimer of the DNA-binding domain from the human oestrogen receptor (ERDBD) and a non-consensus DNA target site in which there is a single base substitution in one half of the palindromic binding site. This substitution results in a 10-fold increase in the dissociation constant of the ERDBD-DNA complex. Comparison of this structure with a structure containing a consensus DNA-binding site determined previously, shows that recognition of the non-consensus sequence is achieved by the rearrangement of a lysine side chain so as to make an alternative base contact. CONCLUSIONS: This study suggests that proteins adapt to recognize different DNA sequences by rearranging side chains at the protein-DNA interface so as to form alternative patterns of intermolecular contacts.

Amino Acid Sequence↗
PubMed1995

Clinical features that differentiate hantavirus pulmonary syndrome from three other acute respiratory illnesses.

To elucidate the early clinical characteristics of hantavirus pulmonary syndrome (HPS), we compared the clinical features of 24 cases of HPS with those of cases of bacteremic pneumococcal pneumonia (n = 30), influenza (n = 33), or unexplained adult respiratory distress syndrome (ARDS, n = 21). On admission, patients with HPS were less likely than outpatients with influenza to have reported sore throat (OR = 0.02, P < .01) and cough (OR = 0.1, P = .01) and were less likely than patients with pneumococcal pneumonia to have lobar infiltrates detected by chest roentgenography (OR = 0, P < .01). Multivariate discriminant analysis revealed that three clinical characteristics at admission (dizziness, nausea or vomiting, and absence of cough) and three initial laboratory abnormalities (low platelet count, low serum bicarbonate level, and elevated hematocrit level) served to identify all patients with HPS and to exclude HPS in at least 80% of patients with unexplained ARDS. These findings warrant further study and should facilitate the early recognition of patients with HPS, who may benefit from early critical-care intervention.

Acute Disease↗
PubMed1994

Promotion of parallel DNA quadruplexes by a yeast telomere binding protein: a circular dichroism study.

Repressor-activator protein 1 (RAP1) has an essential role in the maintenance of yeast telomeres. Yeast telomeric DNA consists of simple repeated G-rich sequences that are bound by RAP1. We have found that RAP1, in addition to its known binding activity for double-stranded DNA, interacts with the G-rich strand containing guanine base (G)-tetrads. We show here using circular dichroism spectroscopy that RAP1 promotes the formation of one particular type of DNA quadruplex, parallel G4-DNA. Furthermore, RAP1 is able to bind to both preformed parallel and antiparallel DNA quadruplexes. These results have implications for the possible use of DNA quadruplexes in telomere-telomere association in vivo.

Animals↗
PubMed1994

Postthaw viability of frozen human ovarian cancer tumor specimens.

Thirty-five human ovarian cancer tumor specimens were frozen by both rapid freeze and slow freeze with dimethyl sulfoxide (DMSO) to determine which method results in optimal postthaw viability. Median viability following a rapid freeze was 8% as compared to 90% with slow freeze with DMSO (P = .0001). Eighty-seven percent of tumors had a viability of > 80% following slow freeze with DMSO. When experimentation on human ovarian cancer tumor cells requires postthaw viability, we recommend slow freeze with DMSO.

Carcinoma↗
PubMed1994

Community-based parentcraft.

Some parents feel unable to ask advice from professionals, or that the advice offered is not appropriate to their own priorities. Lilian Chapman and Peter Jones describe a multi-agency, community-based approach to parentcraft teaching.

Community Health Services↗
PubMed1994

Particle concentration fluorescence immunoassay chemosensitivity assay for human epithelial ovarian cancer.

We adapted particle concentration fluorescence immunoassay technology to develop a new chemosensitivity assay. Six human epithelial ovarian cancer cell lines were incubated with cisplatin for 24 hours. Cells were then labeled with a fluorophore for 30 minutes. Removal of supernatant, washing of tumor cells, and reading of epifluorescence was performed automatically. Percent cell lysis was calculated simultaneously by chromium release assay. The particle concentration fluorescence chemosensitivity assay detected cell lysis in all six cell lines. The chemosensitivity assay detected a larger percent lysis than the chromium release assay in each of the cell lines. The advantages of our chemosensitivity assay are plating efficacy, speed, efficiency, safety, and accuracy.

Cell Death↗
PubMed1993

The crystal structure of a two zinc-finger peptide reveals an extension to the rules for zinc-finger/DNA recognition.

The Cys2-His2 zinc-finger is the most widely occurring DNA-binding motif. The first structure of a zinc-finger/DNA complex revealed a fairly simple mechanism for DNA recognition suggesting that the zinc-finger might represent a candidate template for designing proteins to recognize DNA. Residues at three key positions in an alpha-helical 'reading head' play a dominant role in base-recognition and have been targets for mutagenesis experiments aimed at deriving a recognition code. Here we report the structure of a two zinc-finger DNA-binding domain from the protein Tramtrack complexed with DNA. The amino-terminal zinc-finger and its interaction with DNA illustrate several novel features. These include the use of a serine residue, which is semi-conserved and located outside the three key positions, to make a base contact. Its role in base-recognition correlates with a large, local, protein-induced deformation of the DNA helix at a flexible A-T-A sequence and may give insight into previous mutagenesis experiments. It is apparent from this structure that zinc-finger/DNA recognition is more complex than was originally perceived.

Amino Acid Sequence↗
PubMed1993

The crystal structure of the estrogen receptor DNA-binding domain bound to DNA: how receptors discriminate between their response elements.

The nuclear hormone receptors are a superfamily of ligand-activated DNA-binding transcription factors. We have determined the crystal structure (at 2.4 A) of the fully specific complex between the DNA-binding domain from the estrogen receptor and DNA. The protein binds as a symmetrical dimer to its palindromic binding site consisting of two 6 bp consensus half sites with three intervening base pairs. This structure reveals how the protein recognizes its own half site sequence rather than that of the related glucocorticoid receptor, which differs by only two base pairs. Since all nuclear hormone receptors recognize one or the other of these two consensus half site sequences, this recognition mechanism applies generally to the whole receptor family.

Amino Acid Sequence↗
PubMed1993

Investigation of retroviral involvement in chronic fatigue syndrome.

Within the last few years significant efforts have been made to identify objective reliable diagnostic markers from individuals with chronic fatigue syndrome (CFS). We report the absence of a previously described retroviral marker (HTLV-II gag) in a blinded study of CFS cases. Even with excellent reproducible sensitivities, this marker failed in repeated attempts to distinguish cases from controls. In addition, four other retroviruses (simian T cell leukaemia virus, human spumavirus, bovine leukaemia virus and simian retrovirus) were examined for their presence in these CFS cases and found to be absent. Our findings suggest that these agents, at least as markers, are non-distinguishing for CFS and that other factors may be confounding the resolution of an aetiology to this syndrome.

Biomarkers↗
PubMed1991

Searching: expectant fathers' experiences during labor and birth.

When expectant fathers are present and view labor and birth as a couple experience, they are co-laboring in one of three roles: coach, teammate, or witness. Within these roles are various degrees and types of engagement. Men in the role of coach experience high degrees of physical and mental engagement. Teammates fluctuate between high and low degrees of physical and mental engagement, and witnesses remain at low degrees of engagement until the second stage of labor, when they experience high degrees of mental engagement. The expectant fathers' experience is influenced by the labor guides' activities of gatekeeping, leading, and informing. Men either maintain or redefine their role during labor and birth. Men who experience a sense of not belonging or who are uncomfortable with their role will redefine their role by using the strategy of searching for place. Searching for place involves the steps of identifying an alternative role, engaging in the new role, testing the role, and evaluating the effectiveness of the alternative role. Men who experience a sense of belonging in their new role will maintain this role. May posed the question, "Is it time to fire the coach?" This theory, which requires further testing and validation, does address May's question. Perhaps it is not time to fire the coach, as May indicated, but it is time to provide couples with options in the role expectant fathers play during labor and birth. This theory can guide nurses in relatively risk-free interventions for the expectant father. Nurses can assist expectant fathers in finding a place in labor and birth that will enhance a mutually satisfying birthing experience.

Adult↗