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K C Parker

Publications and source records attributed to K C Parker.

At least 19 recordsLinked to original sources
PubMed2001

Toward a high-throughput approach to quantitative proteomic analysis: expression-dependent protein identification by mass spectrometry.

The isotope-coded affinity tag (ICAT) technology enables the concurrent identification and comparative quantitative analysis of proteins present in biological samples such as cell and tissue extracts and biological fluids by mass spectrometry. The initial implementation of this technology was based on microcapillary chromatography coupled on-line with electrospray ionization tandem mass spectrometry. This implementation lacked the ability to select proteins for identification based on their relative abundance and therefore to focus on differentially expressed proteins. In order to improve the sample throughput of this technology, we have developed a two-step approach that is focused on those proteins for which the abundance changes between samples: First, a new software program for the automated quantification of ICAT reagent labeled peptides analyzed by microcapillary electrospray ionization time-of-flight mass spectrometry determines those peptides that differ in their abundance and second, these peptides are identified by tandem mass spectrometry using an electrospray quadrupole time-of flight mass spectrometer and sequence database searching. Results from the application of this approach to the analysis of differentially expressed proteins secreted from nontumorigenic human prostate epithelial cells and metastatic cancerous human prostate epithelial cells are shown.

Autoanalysis↗
PubMed2001

Eye movement desensitization and reprocessing (EMDR): a meta-analysis.

Eye movement desensitization and reprocessing (EMDR), a controversial treatment suggested for posttraumatic stress disorder (PTSD) and other conditions, was evaluated in a meta-analysis of 34 studies that examined EMDR with a variety of populations and measures. Process and outcome measures were examined separately. and EMDR showed an effect on both when compared with no treatment and with therapies not using exposure to anxiety-provoking stimuli and in pre post EMDR comparisons. However, no significant effect was found when EMDR was compared with other exposure techniques. No incremental effect of eye movements was noted when EMDR was compared with the same procedure without them. R. J. DeRubeis and P. Crits-Christoph (1998) noted that EMDR is a potentially effective treatment for noncombat PTSD. but studies that examined such patient groups did not give clear support to this. In sum, EMDR appears to be no more effective than other exposure techniques, and evidence suggests that the eye movements integral to the treatment, and to its name, are unnecessary.

Adaptation, Psychological↗
PubMed2001

Fine-scale genetic structure in Pinus clausa (Pinaceae) populations: effects of disturbance history.

Spatial autocorrelation analyses of 12 allozyme loci were used to compare genetic structure within populations of two varieties of Pinus clausa. P. clausa var. immuginata populations tend to be uneven-aged, with continuous recruitment in small gaps created by wind damage, whereas P. clausa var. clausa populations are more even-aged, with recruitment postdating periodic canopy fires. Three var. immuginata populations and three matched pairs of var. clausa populations, including both a mature and a nearby recently burned population, were examined. Aggregation of multilocus genotypes at small distances was evident in all young var. clausa populations. Little inbreeding was apparent among juveniles or adults in these populations; their genetic structure is likely to have resulted from limited seed dispersal. Genotypes were not significantly spatially structured in nearby matched mature populations. Genetic structure was less evident in var. immuginata populations. Aggregated genotypes were only apparent in the population where patches included juveniles of similar ages; dense juvenile clumps in the other two var. immuginata populations comprised a variety of ages. Interannual variability in allele frequencies of surviving seedlings may account for the absence of genetic structure in these populations.

Alleles↗
PubMed2000

The murine liver-specific nonclassical MHC class I molecule Q10 binds a classical peptide repertoire.

The biological properties of the nonclassical class I MHC molecules secreted into blood and tissue fluids are not currently understood. To address this issue, we studied the murine Q10 molecule, one of the most abundant, soluble class Ib molecules. Mass spectrometry analyses of hybrid Q10 polypeptides revealed that alpha1alpha2 domains of Q10 associate with 8-9 long peptides similar to the classical class I MHC ligands. Several of the sequenced peptides matched intracellularly synthesized murine proteins. This finding and the observation that the Q10 hybrid assembly is TAP2-dependent supports the notion that Q10 groove is loaded by the classical class I Ag presentation pathway. Peptides eluted from Q10 displayed a binding motif typical of H-2K, D, and L ligands. They carried conserved residues at P2 (Gly), P6 (Leu), and Pomega (Phe/Leu). The role of these residues as anchors/auxiliary anchors was confirmed by Ala substitution experiments. The Q10 peptide repertoire was heterogeneous, with 75% of the groove occupied by a multitude of diverse peptides; however, 25% of the molecules bound a single peptide identical to a region of a TCR V beta-chain. Since this peptide did not display enhanced binding affinity for Q10 nor does its origin and sequence suggest that it is functionally significant, we propose that the nonclassical class I groove of Q10 resembles H-2K, D, and L grooves more than the highly specialized clefts of nonclassical class I Ags such as Qa-1, HLA-E, and M3.

ATP-Binding Cassette Transporters↗
PubMed2000

Attachment security: a meta-analysis of maternal mental health correlates.

This meta-analysis addresses the association between attachment security and each of three maternal mental health correlates. The meta-analysis is based on 35 studies, 39 samples, and 2,064 mother-child pairs. Social-marital support (r = .14; based on 16 studies involving 17 samples and 902 dyads), stress (r = .19; 13 studies, 14 samples, and 768 dyads), and depression (r = .18; 15 studies, 19 samples, and 953 dyads) each proved significantly related to attachment security. All constructs showed substantial variance in effect size. Ecological factors and approach to measuring support may explain the heterogeneity of effect sizes within the social-marital support literature. Effect sizes for stress varied according to the time between assessment of stress and assessment of attachment security. Among studies of depression, clinical samples yielded significantly larger effect sizes than community samples. We discuss these results in terms of measurement issues (specifically, overreliance on self-report inventories) and in terms of the need to study the correlates of change in attachment security, rather than just the correlates of attachment security per se.

Adult↗
PubMed2000

MALDI-TOF based mutation detection using tagged in vitro synthesized peptides.

Matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF) is a powerful method to quickly and accurately determine the masses of peptides. Most genetic analyses, however, begin with PCR amplification of a test sequence to generate DNA, which is more difficult than peptides to analyze by MALDI-TOF. We describe a method that produces a PCR product of any continuous region of coding sequence which can then be used to encode an N-terminally tagged test peptide in a coupled in vitro transcription/translation reaction. The test peptide is purified using the tag, and its mass is measured by MALDI-TOF. Truncations and amino acid substitutions in peptides coded for by the breast cancer susceptibility gene BRCA1 were readily identified using this method. The process can be multiplexed and is amenable to automation, providing an efficient, high-throughput means for mutation discovery and genetic profiling.

Amino Acid Sequence↗
PubMed1999

Relational V-code conditions in a child and adolescent population do warrant treatment.

It is often argued that relational V-code conditions are less serious than classical psychiatric disorders, and that they should therefore receive lower clinical priority or diminished levels of treatment funding. Despite these common assertions, there have been virtually no studies that have used actual case data to evaluate whether such problems are in fact less serious and less worthy of treatment funding. We used actual case data from a universally funded child and family clinic to evaluate these questions. Results showed that both classical diagnoses and relational problems were significantly related to markers of clinical severity. As with previous research, family therapy was not differentially associated with a larger number of treatment sessions.

Adolescent↗
PubMed1998

Genetic analysis on the NifW by utilizing the yeast two-hybrid system revealed that the NifW of Azotobacter vinelandii interacts with the NifZ to form higher-order complexes.

Nitrogenase is a complex metalloenzyme composed of two separately purified proteins designated the Fe-protein and the MoFe-protein. Apart from these two proteins, a number of accessory proteins are essential for the maturation and assembly of nitrogenase. Even though experimental evidence suggests that these accessory proteins are required for nitrogenase activity, the exact roles played by many of these proteins in the functions of nitrogenase are unclear. Our studies were directed to understand the role of two nif accessory proteins, the NifW and the NifZ in the biological nitrogen fixation. To accomplish this, we have utilized a genetic method, the Yeast based Two-Hybrid protein-protein interaction assay. This analysis showed that the NifW could interact with itself to make a multimeric complex. In contrast, the NifZ could not interact with itself. However, the NifZ could interact with the NifW. Previously it was shown that mutating either the NifW or the NifZ have similar effects on the activity of nitrogenase. This observation indicated that both these proteins may exert their regulation on the nitrogenase by a common pathway. Furthermore, it was suggested that the NifW plays a role in the oxygen-protection of the MoFe-protein by direct physical interaction. Our observation that the NifW can interact with itself as well as with the NifZ, suggests that the NifW and the NifZ may form a higher order complex and such a complex may be needed to exert the effects of the NifW or the NifZ on the nitrogenase activity.

Amino Acid Sequence↗
PubMed1998

Identification of yeast proteins from two-dimensional gels: working out spot cross-contamination.

With the complete sequence of the yeast genome now available, efforts by many laboratories are underway to identify each of the spots on two-dimensional (2-D) gels corresponding to the most abundant yeast proteins. The high mass accuracy now attainable using matrix assisted laser desorption/ionization (MALDI)-mass spectrometry equipped with delayed extraction simplifies the process of identification, such that many spots can be unambiguously identified in a short period of time merely by using peptide mass fingerprinting and generally available database matching programs. Although it is not always possible to match spots between gels run by different laboratories, proteins generally yield the same abundant proteolytic fragments when tryptic digestions are performed. Databases containing these signature peptides not only simplify the task of reidentifying proteins from different gels, but also make it possible to identify small amounts of cross-contaminating proteins from different spots, as well as common extraneous contaminants such as human keratins. In this paper, we present data on the identification of > 20 previously unreported yeast proteins from 2-D gels. Some novel proteins were identified from randomly analyzed spots. Focusing on 14 spots in a narrow-pH-range gel, we demonstrate how organizing peak-table data and peptide match-list data into databases enables the identification of a larger percentage of the peaks.

Acrylic Resins↗
PubMed1997

Peptides isolated from HLA-Cw*0304 confer different degrees of protection from natural killer cell-mediated lysis.

HLA class I molecules bind peptides derived from proteins degraded in the cytoplasm and display them for surveillance by the immune system. The recognition of HLA class I molecules by natural killer (NK) cells generally inhibits the lytic process. To investigate the role of peptides in the interaction between HLA class I molecules and NK receptors, we first had to identify representative endogenous peptides. Individual peptides bound to HLA-Cw*0304 were isolated and sequenced by tandem mass spectrometry. These peptides ranged in length from 8 to 11 residues and shared an alanine at position 2 and a C-terminal leucine. The murine transporters associated with antigen processing (TAP)-deficient cell line RMA-S was transfected with HLA-Cw*0304 to test whether HLA molecules loaded with a single peptide could deliver the inhibitory signal to NK cells expressing p58.2, which is a killer cell inhibitory receptor known to interact with HLA molecules bearing the HLA-Cw3 public epitope. We found that, in the absence of exogenous peptides, the HLA-Cw*0304 transfectants were killed at levels comparable to untransfected RMA-S cells whereas protection from lysis required both HLA-Cw*0304 heavy chain expression and an exogenously added HLA-Cw*0304-binding peptide. Importantly, not only were HLA-Cw*0304-binding peptides required for protection, but the ability of individual peptides to provide protection differed widely. These studies indicate that the ability to distinguish between subsets of peptides may be a general feature of HLA class I recognition by NK cells.

Amino Acid Sequence↗
PubMed1997

Identification of an epitope derived from human proteolipid protein that can induce autoreactive CD8+ cytotoxic T lymphocytes restricted by HLA-A3: evidence for cross-reactivity with an environmental microorganism.

The demyelination process that occurs in the central nervous system (CNS) of patients with multiple sclerosis (MS) is in part due to an inflammatory response in which CD4+ and CD8+ T cells and macrophages infiltrate white matter. In this study, we have identified a peptide sequence derived from the CNS-specific myelin protein proteolipid protein (PLP) which could bind to HLA-A3 and induce a HLA-A3-restricted CD8+ CTL response from HLA-A3+ donors. These PLP peptide-specific CTL could lyse HLA-A3+ target cells pulsed with a homologous peptide derived from the CRM1 protein of Saccharomyces cerevisae. These findings demonstrate the immunogenic potential of a PLP-derived peptide for generation of autoreactive HLA-A3-restricted CD8+ CTL, and further show that these CTL can be activated by a peptide derived from a common environmental microorganism.

Adult↗
PubMed1997

Mothers' representations of their infants assessed prenatally: stability and association with infants' attachment classifications.

The stability and predictive validity of classifications of mothers' representations of their infants as determined by the Working Model of the Child Interview (WMCI) were examined. Concordance between mothers' representations of their infants assessed prenatally and again one year later and infant Strange Situation (SS) attachment classifications at 12 months was also examined. WMCI classifications were stable over 12 months in 80% of mothers, compared to 51% expected by chance alone. Pregnancy WMCIs predicted infant SS classifications in 74% of cases, compared to 54% expected by chance. Concordance between 11-month WMCI and 12-months SS classifications was 73% (vs. 55% expected by chance). Problems with the skewed distribution of the sample, the low concordance between pregnancy and 11 months for one of the three classifications, and future directions for research are discussed.

Female↗
PubMed1996

Peptide sequence requirements for the recognition of HLA-B*2705 by specific natural killer cells.

NK clones that were inhibited by target cell expression of HLA-B*2705 displayed peptide-specific recognition of HLA-B*2705. To evaluate the specificity of this recognition, synthetic versions of 14 endogenous ligands of HLA-B*2705 were tested for their ability to provide protection from NK-mediated lysis by binding to surface HLA-B*2705 molecules on RMA-S cells deficient in the transporter for Ag presentation. Several unrelated peptides inhibited lysis by the same NK clones. Despite similar capacities to stabilize HLA-B*2705 molecules on RMA-S cells, the 14 peptides differed widely in their abilities to provide protection. Single amino acid substitutions in both a protective and a nonprotective peptide revealed the importance of residues 7 and 8 in the peptide for recognition by NK clones, thus localizing the peptide influence to a polymorphic region of the alpha-helix of HLA class I molecules known to control discrimination among allelic variants of HLA-B and HLA-C by NK cells.

ATP Binding Cassette Transporter, Subfamily B, Mem↗
PubMed1996

Molecular analysis of presentation by HLA-A2.1 of a promiscuously binding V3 loop peptide from the HIV-envelope protein to human cytotoxic T lymphocytes.

P18(IIIB) is a highly immunogenic peptide from the V3 loop of the HIV-1 gp160 envelope protein that is presented promiscuously by multiple class I MHC molecules. Understanding the molecular basis for promiscuous presentation may have many practical applications. As the highly prevalent HLA-A2.1 class I molecule is known to present P18(IIIB) for recognition by cytotoxic T lymphocytes (CTL) found in peripheral blood mononuclear cells of HIV+ donors, a P18(IIIB)-specific CTL line was generated from and HLA-A2(+), HIV- donor in order to define the molecular basis for, and ultimately improve upon the binding of, this peptide to HLA-A2.1. The minimal epitope recognized by the line was a decamer, I10, with the sequence RGPGRAFVTI. Interestingly, this decamer is identical to the minimal epitope from P18(IIIB) seen by murine CTL restricted by H-2Dd. A panel of Ala-substituted peptides was employed in MHC-binding and T cell response studies to identify MHC- and TCR-binding residues. Notably, many of the agretopic and epitopic residues identified were identical to those involved in the corresponding interactions of I10 with the H-2Dd MHC molecule and murine I10-specific CTL. The I10 peptide does not contain the described HLA-A2.1 binding motif. Instead a Pro at P3, a Phe at P7 and an Ile at P10 are utilized for MHC binding. Agretopic residue similarities with the hepatitis B nucleocapsid decamer suggest that these residues may comprise an alternative motif of anchors utilized by decamers for binding to HLA-A2.1.

Amino Acid Sequence↗
PubMed1995

Irradiated sporozoite vaccine induces HLA-B8-restricted cytotoxic T lymphocyte responses against two overlapping epitopes of the Plasmodium falciparum sporozoite surface protein 2.

Vaccines designed to protect against malaria by inducing CD8+ cytotoxic T lymphocytes (CTL) in individuals of diverse HLA backgrounds must contain multiple conserved epitopes from various preerythrocytic-stage antigens. Plasmodium falciparum sporozoite surface protein 2 (PfSSP2) is considered an important antigen for inclusion in such vaccines, because CD8+ CTL against the P. yoelii SSP2 protect mice against malaria by eliminating infected hepatocytes. To develop PfSSP2 as a component of malaria vaccines, we investigated the presence of anti-PfSSP2 CTL in two HLA-B8+ volunteers immunized with irradiated P. falciparum sporozoites and characterized their CTL responses using PfSSP2-derived 15-amino acid peptides bearing the HLA-B8-binding motif. Peripheral blood mononuclear cells from both volunteers stimulated with recombinant vaccinia expressing PfSSP2 displayed antigen-specific, genetically restricted, CD8+ T cell-dependent CTL activity against autologous target cells expressing PfSSP2. Of the five HLA-B8 motif-bearing 15-mers identified in the PfSSP2 sequence, two peptides sharing a 10-amino acid overlap sensitized HLA-B8-matched target cells from both volunteers for lysis by peptide-stimulated effectors. The CTL activity was HLA-B8 restricted and dependent on CD8+ T cells. Analysis of the three shorter peptides representing HLA-B8 motif-bearing sequences within the two positive peptides for their ability to bind to HLA-B8 in vitro, and to sensitize target cells for lysis by effectors stimulated with the 15-mers, identified two overlapping HLA-B8-restricted CTL epitopes. Available data indicate that the sequence of one CTL epitope is conserved and the other is variant among P. falciparum isolates. Circulating activated CTL against the conserved epitope could be directly identified in one of the two volunteers. The identification of two HLA-B8-restricted CTL epitopes on PfSSP2 provides data critical to developing an epitope-based anti-liver stage malaria vaccine.

Amino Acid Sequence↗
PubMed1995

Identification of the peptide binding motif for HLA-B44, one of the most common HLA-B alleles in the Caucasian population.

Most peptides that bind to a particular MHC class I molecule share amino acid residues that are thought to physically "anchor" the peptide to polymorphic pockets within the class I binding site. Sequence analysis of endogenous peptides bound to HLA-B44 revealed two potential dominant anchor residues: Glu at P2 and Tyr, or occasionally Phe, at P9. In vitro assembly assays employing synthetic peptides and recombinant HLA-B44 produced by Escherichia coli revealed that an acidic amino acid at P2 was necessary for promoting stable peptide binding to HLA-B44. Surprisingly, although Tyr was almost exclusively found at P9 of the endogenous peptide sequences, a wide variety of amino acid residues such as Leu, Ala, Arg, Lys, His, and Phe could be tolerated at this position. Using this information, we identified antigenic peptides from the influenza virus components nonstructural protein 1 and nucleoprotein that are presented by HLA-B44 to antiinfluenza type A cytotoxic T lymphocytes. In addition, cytotoxic T lymphocytes induced by these antigenic peptides were shown to be capable of recognizing endogenously processed peptides from influenza-infected cells, indicating a potential use for these peptides in vaccine development. Finally, molecular models were created to investigate the possible ways in which the anchor residues might function to stabilize the binding of peptides to HLA-B44, and these models indicate that the acidic residue at P2 most likely interacts primarily with Lys 45 of the HLA-B44 heavy chain and makes additional contacts with Ser 67 and Tyr 9.

Alleles↗