Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: UNIPROT:P06889 (Mol)
 630,302 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Two malate dehydrogenase isoforms, named MDH1 and MDH2, have been purified to homogeneity from Trypanosoma cruzi epimastigotes. Both enzymes consist of subunits with a molecular mass close to 33 kDa; native molecular mass determination by gel filtration, however, indicated that MDH1 is a dimer, whereas MDH2 is a tetramer. Both isoforms did not cross-react immunologically. The N-termini of both MDH isoforms and several tryptic peptides of MDH1 (amounting to about one third of the complete molecule) have been sequenced by automated Edman degradation. The tryptic digests of both enzymes have also been analysed by mass spectrometry (MALDI-TOF MS). The apparent Km values in both directions of the reaction have been determined, as well as the possible inhibition by excess of the substrate oxaloacetate. The sequence data, together with the pI values and the presence or absence of oxaloacetate inhibition indicate that the dimeric MDH1 is the mitochondrial isoenzyme, whereas the tetrameric MDH2 is the glycosomal isoenzyme. No evidence was found for the presence of a cytosolic isoform.
Mol Biochem Parasitol 2000 Feb 05
PMID:Tetrameric and dimeric malate dehydrogenase isoenzymes in Trypanosoma cruzi epimastigotes. 1069 43

Genomic imprinting is the result of a gamete-specific modification leading to parental origin-specific gene expression in somatic cells of the offspring. Several embryonal tumors show loss of imprinting of genes clustered in human chromosome 11p15.5, an important tumor suppressor gene region, harboring several normally imprinted genes. TSSC3, a gene homologous to mouse TDAG51, implicated in Fas-mediated apoptosis, is also located in this region between hNAP2 and p57 (KIP2). TSSC3 is the first apoptosis-related gene found to be imprinted in placenta, liver and fetal tissues where it is expressed from the maternal allele in normal human development. This study investigated the imprinting status of TSSC3 in human normal, adult brain and in human neuroblastomas, medulloblastomas and glioblastomas. A polymorphism in exon 1 at position 54 was used to analyze the allelic expression of the TSSC3 gene by a primer oligo base extension (PROBE) assay using matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF MS). We found that the TSSC3 gene is not imprinted in human normal, adult brain and blood. In contrast, strong allelic bias resembling imprinting could be detected in most examined tumor specimens. The results demonstrate for the first time that the tumors under investigation are associated with a retention of imprinting of a potential growth inhibitory gene.
Hum Mol Genet 2000 Mar 22
PMID:Retention of imprinting of the human apoptosis-related gene TSSC3 in human brain tumors. 1074 82

Biomolecular interaction analysis mass spectrometry (BIA/MS) is a multiplexed analytical technique that utilizes a unique combination of surface plasmon resonance (SPR) and matrix assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) for the detection and analysis of small amounts of proteins residing in complex biological systems. In order to achieve high sensitivity during BIA/MS, certain experimental parameters and sequences of events need to be optimized and maintained. Immobilized ligand density, flow rate and biosensor control (in SPR-BIA) and matrix choice and application (in MALDI-TOF MS) have significant influence on the final outcome of the BIA/MS analysis and, consequently, need to be optimized and carefully controlled. In addition, chip washing and cutting are essential in converting the SPR-active sensor chips into target surfaces amenable to MALDI-TOF MS. Reviewed here are the prerequisites for successfully interfacing SPR-BIA with MALDI-TOF MS.
J Mol Recognit
PMID:Practical considerations in BIA/MS: optimizing the biosensor-mass spectrometry interface. 1086 9

AP-2 is a cell-type specific, developmentally regulated transcription factor which has been described as a critical regulator of gene expression during vertebrate development and embryogenesis. Although the overall domains of this factor necessary for their activity have been identified, the exact identity of AP-2 amino acid residues responsible for its interaction with the DNA structure has not yet been described. Here, we describe the identification of a region of AP-2 which was protected by an oligonucleotide probe containing its binding site from trypsin digestion, monitored by peptide mapping by MALDI-TOF mass spectrometry. Furthermore, we analyzed the relative in vitro DNA-binding activity, the stimulatory potency on the AP-2-dependent APOE promoter, as well as the ability to inhibit the effect of the wild-type protein of each one of a set of single-site substitution AP-2 mutants spanning the identified region. Taken together, our data clearly demonstrate that the region between amino acid residues 252-260 of AP-2 is essential for its DNA-binding activity. Particularly, the individual substitution in any of the residues 253, 254, 255, 257 or 260 is sufficient for completely abolishing the interaction with DNA and the stimulation of APOE promoter activity. These results indicate a crucial role of this region in the formation of an active DNA-binding domain and strongly suggest that these residues provide direct contacts with the DNA structure at the AP-2 binding site.
J Mol Biol 2000 Aug 25
PMID:Identification of amino acid residues of transcription factor AP-2 involved in DNA binding. 1096 87

[His(7)]-corazonin has recently been identified in the corpora cardiaca (CC) of two locust species, the migratory locust, Locusta migratoria and the desert locust, Schistocerca gregaria, as the dark colour inducing neurohormone. Here, we investigate whether [His(7)]-corazonin occurs in the brain-CC axis of a Schistocerca albino strain. From data obtained by immunocytochemistry, injection experiments, chromatographic and mass spectrometric analysis of brain and CC tissues, it could be concluded that an albino strain of S. gregaria from Denmark contains authentic [His(7)]-corazonin. This was unequivocally demonstrated by sequencing the [His(7)]-corazonin-immunoreactive factor in albino Schistocerca brain-CC extracts with ESI-Qq-oa-TOF mass spectrometry. Albinism in this strain is hence not caused by the deficiency of authentic [His(7)]-corazonin in the brain-CC axis, nor by defects in release. Conversely to L. migratoria albinos, injection of [His(7)]-corazonin failed to induce dark pigmentation in Schistocerca albinos. Therefore, albinism in the investigated Schistocerca strain is likely to be situated at the level of the receptor, signal transduction mechanisms or of pigment biosynthesis.
Mol Cell Endocrinol 2000 Oct 25
PMID:The pigmentotropic hormone [His(7)]-corazonin, absent in a Locusta migratoria albino strain, occurs in an albino strain of Schistocerca gregaria. 1106 56

For various diagnostic analyses and the studies of functional genomics, the use of an accurate and cost-effective analytic platform to analyze large numbers of samples is essential. An automated platform called MassArray (Sequenom, Inc, San Diego, CA), designed for high-throughput diagnostic analyses, has recently been validated. The platform combines miniaturized, two-dimensional chip arrays with proven high-fidelity enzymatic procedures and matrix-assisted laser-desorption/ionization time-of-flight (MALDI-TOF) mass spectrometry. Nanoliter dispensing of samples in high-density formats of 384 or greater results in improved throughput and reduced costs. Automation prevails from the initial assay design through sample processing and data analysis, for the most part eliminating the labor component of assay development and implementation. The MassArray platform is being used in the following areas: (1) molecular diagnosis of genetic disease and infectious agents, (2) pharmacogenomics, (3) paternity and/or identity testing, and (4) agriculture (e.g., marker-assisted breeding). MALDI-TOF mass spectrometry can also be used for analyzing proteins; therefore, genotype/phenotype testing can be performed on a single platform.
Mol Diagn 2000 Dec
PMID:Automated mass spectrometry: a revolutionary technology for clinical diagnostics. 1117 98

The kinetics of solvent accessibility at the protein-protein interface between thrombin and a fragment of thrombomodulin, TMEGF45, have been monitored by amide hydrogen/deuterium (H/2H) exchange detected by MALDI-TOF mass spectrometry. The interaction is rapid and reversible, requiring development of theory and experimental methods to distinguish H/2H exchange due to solvent accessibility at the interface from H/2H exchange due to complex dissociation. Association and dissociation rate constants were measured by surface plasmon resonance and amide H/2H exchange rates were measured at different pH values and concentrations of TMEGF45. When essentially 100% of the thrombin was bound to TMEGF45, two segments of thrombin became completely solvent-inaccessible, as evidenced by the pH insensitivity of the amide H/2H exchange rates. These segments form part of anion-binding exosite I and contain the residues for which alanine substitution abolishes TM binding. Several other regions of thrombin showed slowing of amide exchange upon TMEGF45 binding, but the exchange remained pH-dependent, suggesting that these regions of thrombin were rendered only partially solvent-inaccessible by TMEGF45 binding. These partially inaccessible regions of thrombin form both surface and buried contacts into the active site of thrombin and contain residues implicated in allosteric changes in thrombin upon TM binding.
J Mol Biol 2001 Feb 23
PMID:Solvent accessibility of the thrombin-thrombomodulin interface. 1117 15

The cyanobacterium Synechocystis sp. PCC 6803 is an ideal model organism for the proteome study of light-induced gene expression because the whole genomic sequence has been determined. The soluble proteins extracted from light- and dark-cultured cells were separated by two-dimensional polyacrylamide gel electrophoresis. Light-induced protein spots electroblotted on a polyvinyldiene difluoride membrane were analyzed by N-terminal Edman sequence determination and followed by CyanoBase. The tryptic digests of some proteins were also confirmed by matrix-assisted laser desorption ionization/time-of-flight (MALDI-TOF) and MS-Fit search. Interestingly, eight proteins were related to photosynthesis and respiration (RbcS/L, CbbA, Gap2, AtpB, CpcB, PsbO, and PsbU). Four proteins (SodB, DnaK, GroEL2, and Tig) were involved in cellular processes and the functions of another two proteins (rehydrin and membrane protein) were unknown. The proteome analysis by N-terminal Edman sequencing and MALDI-TOF enabled us to characterize one-shot protein profiles expressed under different physiological conditions.
Mol Cells 2000 Dec 31
PMID:Proteome analysis of light-induced proteins in Synechocystis sp. PCC 6803: identification of proteins separated by 2D-PAGE using N-terminal sequencing and MALDI-TOF MS. 1121 77

CE fractions may also be collected and then subjected to additional analysis. Nanoliter fractions containing size or shape fractionated DNA fragments can be collected on moving affinity membranes (125) or into sample chambers (126). The exact timing of the collection steps is achieved by determining the velocity of each individual zone measured between two detection points near the end of the capillary. The DNA samples may subsequently be identified by probe hybridization, or by PCR-linked sequencing. Capillary fractions containing metabolites and derivatives of DNA and small DNA adducts can also be sampled, and then characterized directly by highly sensitive MALDI-TOF atomic analysis (112-118) and ESI-MS (118,119). The automation and integration of PCR and CE analysis (PCR-CE) on a microchip (3-12,96) will also contribute greatly to its adoption as the analysis tool of choice. Significantly, these tools will be applied for DNA sequencing (75,108), for genome mapping (65) and genotyping (42-46), for improved certainty in disease detection (3-6,107,120) and for DNA mutation analysis (2-12,27,58). Recent improvements in the design CAE arrays and associated equipment such as the radial CAE microplate and rotary confocal signal detection system (127) overcome some of the detection limitations of linear CAE and microchip devices and allow the parallel genotyping of 96 samples in about 120 s. The integration of microreactive capillary surface assays (128) and "in-capillary" analysis will also lead to further increases in the speed and sensitivity of CE-based analysis. The recent announcement of the completion of the first draft sequence of the 90% of the entire human genome within 6 mo by Celera Genomics by sequencing random DNA fragments using several hundred ABI 3700 machines (129) illustrates the enormous efficiency realized through the automation of DNA sequencing by CAE. Sequencing was performed at an average rate of approximately 6 x 10(9) bases/yr. The CAE machines will now be employed for a concerted resequencing of genome elements to create an extremely high-density polymorphism map of the entire genome (130). This map will be based principally on single nucleotide polymorphisms, and will catapult human medicine into a new era of closely detailed genetic trait mapping to identify the genetic basis of multi-gene diseases.
Methods Mol Biol 2001
PMID:The application of capillary electrophoresis for DNA polymorphism analysis. 1121 41

Juvenile hormone esterase (JHE, EC 3.1.1.1) from whole Drosophila melanogaster prepupae has previously been purified by selective precipitations, isoelectric focussing and two column chromatography steps. JHE bands from dried silver-stained SDS-PAGE gels of that material were digested with trypsin. The masses of the tryptic digest peptides were determined by MALDI-TOF mass spectrometry. Only one predicted gene product (CG8425) from the D. melanogaster genome matches the JHE tryptic fingerprint with high confidence. This predicted JHE sequence includes features that are conserved among all active members of the serine carboxylesterase multigene family as well as features peculiar to JHEs from other species. Also we show that this JHE can be purified by an alternative method using anion exchange chromotography followed by trifluoromethylketone affinity chromatography. A cDNA encoding this JHE was isolated using 3' and 5' RACE. This sequence is in agreement with the Drosophila genome project's prediction except that the sixth predicted intron is not removed; instead there is a stop codon followed by a polyadenylation signal and a polyA tail.
Insect Biochem Mol Biol 2001 Apr 27
PMID:Identification of a juvenile hormone esterase gene by matching its peptide mass fingerprint with a sequence from the Drosophila genome project. 1126 90


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