Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: UMLS:C0016632 (Fox)
 1,461 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

A full-length clone of pig kidney fructose 1,6-bisphosphatase (D-fructose-1,6-bisphosphate 1-phosphohydrolase, EC 3.1.3.11) was isolated by screening a cDNA library for complementation of an Escherichia coli fbp deletion mutation. The open reading frame of 1011 bases corresponds to 337 amino acids, two more than have been previously reported [Marcus, F., Edelstein, I., Reardon, I. & Heinrikson, R. L. (1982) Proc. Natl. Acad. Sci. USA 79, 7161-7165]. The extra two amino acids (Ala-Lys) are located at the C-terminal end of the protein as an extension. Comparison of the deduced amino acid sequence with the reported (see above) and revised amino acid sequence [Harrsch, P. B., Kim, Y., Fox, J. L. & Marcus, F. (1985) Biochem. Biophys. Res. Commun. 133, 520-526] indicates three differences in addition to the C-terminal extension. Gln-20, Thr-96, and Asn-199 in the amino acid sequence are found to be Glu, Ser, and Asp, respectively. Since the x-ray structure of the pig kidney enzyme has been reported, the cDNA clone will allow the construction of site-specific mutants to help test possible structure-function relationships in this important metabolic enzyme.
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PMID:Isolation and sequence analysis of the cDNA for pig kidney fructose 1,6-bisphosphatase. 131 79

The gene for glycoprotein gB2 of herpes simplex virus type 2 strain 333 was cloned, sequenced, and expressed in mammalian cells. The gB2 protein had an overall nucleotide and amino acid sequence homology of 86% with the cognate gB1 protein. However, of the 125 amino acid substitutions or deletions, only 12.5% were conservative replacements. These differences were clustered within an NH2-terminal region, a central region, and a COOH-terminal region, resulting in domains of near identity broken by small regions of marked divergence. Regions of greatest homology included a 90-amino-acid stretch starting at residue 484 and 39 amino acids spanning residues 835 to 873, which cover a rate-of-entry locus mapped to Ala-552 and a syn locus mapped to Arg-857, respectively, in gB1 by Bzik et al. (D. J. Bzik, B. A. Fox, N. A. DeLuca, and S. Person, Virology 133:301-314, 1984). Pellett et al. (P. E. Pellett, K. G. Kousoulas, L. Pereira, and B. Roizman, J. Virol. 53:243-253, 1985) mapped the mutations in three monoclonal antibody-resistant gB1 mutants between amino acids 273 and 443. These epitopes are included in a region of 98 residues identical between gB1 and gB2. The identity of this protein was verified by placing a truncated gene lacking the 303 carboxyl-terminal amino acids of gB2 into mammalian COS and CHO cells. Expression was demonstrated by immunofluorescence and radioimmunoprecipitation. This protein will be purified from the stable CHO cell lines and compared with gB1 for immunogenicity and protective efficacy in animal challenge models.
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PMID:Structure and expression of the herpes simplex virus type 2 glycoprotein gB gene. 302 64

To check and clarify existing data on receptor-interacting residues in the human C5a anaphylatoxin, we tested mutant C5a proteins obtained by site-directed mutagenesis of a recombinant human C5a (rhC5a) cDNA clone for structural and functional integrity. Amino acid positions in three different regions of the molecule were investigated: Arg74 at the C-terminus, Arg40 and Pro45 located in the core region, and Lys14 and Lys19, Lys20 in the N-terminus. Des-Arg74-rhC5a displayed only a residual 3-4% functional activity in the myeloperoxidase-release assay from human granulocytes while retaining the three-dimensional solution structure of wild-type (wt)-rhC5a as shown by circular dichroism (CD) spectroscopy. Des-Arg74-rhC5a was able to activate the human C5a receptor transiently expressed in Xenopus oocytes, but was inactive in the heterologous guinea pig (gp) ileum-contraction assay. These results reveal profound differences between the guinea pig and human C5a-receptor ligand-binding characteristics. Exchange of the core residue Arg40 by a glycine did not significantly affect functional C5a activity, in contrast to a previous observation [Mollison, K. W., Mandecki, W., Zuiderweg, E. P., Fayer, L., Fey, T. A., Krause, R. A., Conway, R. G., Miller, L., Edalji, R. P., Shallcross, M. A., Lane, B., Fox, J. L., Greer, J. & Carter, G. W. (1989) Identification of receptor-interacting residues in the inflammatory complement protein C5a by site-directed mutagenesis, Proc. Natl Acad. Sci. USA 86, 292-296], nor did exchange of the conserved Pro45 residue by the C3a analogue glutamic acid, a mutation expected to alter the whole geometry of the loop connecting helix III-helix IV (including Arg40) of the C5a molecule. Thus, participation of this loop in receptor interaction appears unlikely. While exchange of the N-terminal Lys14 residue by alanine did not significantly affect functional activity, a double replacement of Lys19 and Lys20 by alanine residues reduced activity more than 30-fold. These results confirm Lys19 and/or Lys20 as a putative receptor-interacting site, although we could not obtain a CD spectrum of this important mutant due to poor expression.
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PMID:Site-specific mutagenesis of residues in the human C5a anaphylatoxin which are involved in possible interaction with the C5a receptor. 811 41

The glycoprotein (GP) Ib-IX-V complex constitutively expressed on the platelet plasma membrane mediates initial adhesion of circulating platelets to vessel wall matrix at high shear, and shear-induced platelet aggregation. In both cases, this involves binding of GP Ib-IX-V to the adhesive glycoprotein, von Willebrand Factor (vWF). vWF binding to GP Ib-IX-V rapidly induces platelet activation, leading to cytoskeletal rearrangement, shape change, and secretion that enables alphaIIbbeta3 integrin (GP IIb-IIIa)-dependent platelet aggregation. All these events are critical in (patho)physiological thrombus formation. The recent discovery that the signaling protein, 14-3-3 zeta, copurifies with the GP Ib-IX complex (minus GP V) [Du, X., Harris, S. J., Tetaz, T. J., Ginsberg, M. H., & Berndt, M. C. (1994) J. Biol. Chem. 269, 18287-18290] indicated a potential mechanism for vWF-dependent signaling. The aim of the present study was to identify discrete amino acid sequences that bind 14-3-3 zeta within the cytoplasmic domain of the receptor. As an initial screening assay, overlapping synthetic peptides based on the cytoplasmic domains of GP Ibalpha (100 residues), GP Ibbeta (34 residues), GP IX (5 residues), and GP V (16 residues) were immobilized and assessed for the ability to bind purified 14-3-3 zeta. The C-terminal sequence GHSL of GP Ibalpha was identified as one 14-3-3 zeta interactive sequence, consistent with previous results [Du, X., Fox, J. E., & Pei, S. (1996) J. Biol. Chem. 271, 7362-7367]. Binding of 125I-labeled 14-3-3 zeta to GHSL-containing peptides was inhibitable by unlabeled 14-3-3 zeta and by anti-14-3-3 zeta IgG. Ala-walking through the GHSL sequence suggested all residues were necessary for optimal binding. In addition, 14-3-3 zeta bound with lower affinity to a peptide based on the central region of the GP Ibalpha cytoplasmic domain (Arg-557-Gly-575), whereas peptide sequences within the cytoplasmic domains of GP Ibbeta (Arg-160-Arg-175) and GP V (Lys-529-Gly-544) bound 14-3-3 zeta with comparable affinity to the GHSL-containing peptide. Soluble GHSL-containing peptides, GP Ibbeta- and GP V-based peptides semidissociated 14-3-3 zeta from GP Ib-IX-V or GP Ib-IX in platelet extracts as analyzed by immunoprecipitation, suggesting these sequences, at least partially, mediate the GP Ib-IX-V-14-3-3 zeta interaction in cells. Further, phosphorylation of the GP Ibbeta peptide at a site corresponding to a protein kinase A phosphorylation site (Ser-166) enhanced the affinity of 14-3-3 zeta binding by approximately 8-fold, suggesting phosphorylation as a potential mechanism for regulating 14-3-3 zeta association with the GP Ib-IX-V complex.
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PMID:Binding of purified 14-3-3 zeta signaling protein to discrete amino acid sequences within the cytoplasmic domain of the platelet membrane glycoprotein Ib-IX-V complex. 942 86

T4MOC is a 12.3 kDa soluble Rieske ferredoxin that is obligately required for electron transfer between the oxidoreductase and diiron hydroxylase components of toluene 4-monooxygenase from Pseudomonas mendocina KR1. Our preliminary 1H NMR studies of oxidized and reduced T4MOC [Markley, J. L., Xia, B., Chae, Y. K., Cheng, H., Westler, W. M., Pikus, J. D., and Fox, B. G. (1996) in Protein Structure Function Relationships (Zaidi, Z., and Smith, D., Eds.) pp 135-146, Plenum Press, London] revealed the presence of hyperfine-shifted 1H resonances whose short relaxation times made it impractical to use nuclear Overhauser effect (NOE) measurements for assignment purposes. We report here the use of selective isotopic labeling to analyze the hyperfine-shifted 1H, 2H, and 15N signals from T4MOC. Selective deuteration led to identification of signals from the four Hbeta atoms of cluster ligands C45 and C64 in the oxidized and reduced forms of T4MOC. In the reduced state, the Curie temperature dependence of the Hbeta protons corresponded to that predicted from the simple vector spin-coupling model for nuclei associated with the localized ferric site. The signal at 25.5 ppm in the 1H spectrum of reduced T4MOC was assigned on the basis of selective 2H labeling to the His Hepsilon1 atom of one of the cluster ligands (H47 or H67). This assignment was corroborated by a one bond 1H-13C correlation (at 25.39 ppm 1H and 136.11 ppm 13C) observed in spectra of [U-13C]T4MOC with a 1H-13C coupling constant of approximately 192 Hz. The carbon chemical shift and one bond coupling constant are those expected for 1Hepsilon1-13Cepsilon1 in the imidazolium ring of histidine and are inconsistent with values expected for cysteine 1Halpha-13Calpha. The His Hepsilon1 proton exhibited weak Curie temperature dependence from 283 to 303 K, contrary to the anti-Curie temperature dependence predicted from the spin coupling model for nuclei associated with the localized ferrous site. A 1H peak at -12.3 ppm was observed in spectra of reduced T4MOC; this signal was found to correspond to a hydrogen (probably in an H-bond to the cluster) that exchanged with solvent with a half-time of about 2 days in the oxidized state but with a much longer (undetectable) half-time in the reduced state. These results with T4MOC call into question certain 1H assignments recently reported on the basis of NOE measurements for the comparable Rieske ferredoxin component of an evolutionarily related alkene monooxygenase from Xanthobacter sp. Py2 [Holz, R. C., Small, F. J., and Ensign, S. A, (1997) Biochemistry 36, 14690-14696]. Selective 15N labeling was used to identify hyperfine-shifted 15N NMR signals from the backbone nitrogens of all four cluster ligands (C45, H47, C64, and H67), from the Nepsilon2 atoms of the two histidine ligands (H47 and H67), and from nonligand Gln and Ala residues (Q48 and A66) present in the cluster-binding motif of T4MOC in the oxidized and reduced states. The results indicate that the Ndelta1 of each of the two ligand histidines of T4MOC are ligated to an iron atom and reveal a pattern of H-bonding to the Rieske [2Fe-2S] center involving four (H47, Q48, A66, and H67 of T4MOC) of the five backbone amide H-bonds expected on the basis of comparison with the crystal structures of other related Rieske proteins; the fifth backbone amide (I50 of T4MOC) failed to exhibit a hyperfine shift. This anomaly may arise from the lack of an associated disulfide in T4MOC, a fundamental structural difference between the three types of Rieske proteins that may be related to functional diversity in this protein family.
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PMID:Detection and classification of hyperfine-shifted 1H, 2H, and 15N resonances of the Rieske ferredoxin component of toluene 4-monooxygenase. 988 13

Threonine 37 is conserved among all the members of the old yellow enzyme (OYE) family. The hydroxyl group of this residue forms a hydrogen bond with the C-4 oxygen atom of the FMN reaction center of the enzyme [Fox, K. M. & Karplus, P. A. (1994) Structure 2, 1089-1105]. The position of Thr-37 and its interaction with flavin allow for speculations about its role in enzyme activity. This residue was mutated to alanine and the mutant enzyme was studied and compared with the wild-type OYE1 to evaluate its mechanistic function. The mutation has different effects on the two separate half-reactions of the enzyme. The mutant enzyme has enhanced activity in the oxidative half-reaction but the reductive half-reaction is slowed down by more than one order of magnitude. The peaks of the absorption spectra for enzyme bound with phenolic compounds are shifted toward shorter wavelengths than those of wild-type OYE1, consistent with its lower redox potential. It is suggested that Thr-37 in the wild-type OYE1 increases the redox potential of the enzyme by stabilizing the negative charge of the reduced flavin through hydrogen bonding with it.
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PMID:The role of threonine 37 in flavin reactivity of the old yellow enzyme. 1009 75

The conformation of the 20-residue antibiotic ionophore alamethicin in macroscopically oriented phospholipid bilayers has been studied using (15)N solid-state nuclear magnetic resonance (NMR) spectroscopy in combination with molecular modeling and molecular dynamics simulations. Differently (15)N-labeled variants of alamethicin and an analog with three of the alpha-amino-isobutyric acid residues replaced by alanines have been investigated to establish experimental structural constraints and determine the orientation of alamethicin in hydrated phospholipid (dimyristoylphosphatidylcholine) bilayers and to investigate the potential for a major kink in the region of the central Pro(14) residue. From the anisotropic (15)N chemical shifts and (1)H-(15)N dipolar couplings determined for alamethicin with (15)N-labeling on the Ala(6), Val(9), and Val(15) residues and incorporated into phospholipid bilayer with a peptide:lipid molar ratio of 1:8, we deduce that alamethicin has a largely linear alpha-helical structure spanning the membrane with the molecular axis tilted by 10-20 degrees relative to the bilayer normal. In particular, we find compatibility with a straight alpha-helix tilted by 17 degrees and a slightly kinked molecular dynamics structure tilted by 11 degrees relative to the bilayer normal. In contrast, the structural constraints derived by solid-state NMR appear not to be compatible with any of several model structures crossing the membrane with vanishing tilt angle or the earlier reported x-ray diffraction structure (Fox and Richards, Nature. 300:325-330, 1982). The solid-state NMR-compatible structures may support the formation of a left-handed and parallel multimeric ion channel.
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PMID:Conformation of alamethicin in oriented phospholipid bilayers determined by (15)N solid-state nuclear magnetic resonance. 1150 81

Forkhead box P (FoxP) proteins are members of the versatile Fox transcription factors, which control the timing and expression of multiple genes for eukaryotic cell homeostasis. Compared to other Fox proteins, they can form domain-swapped dimers through their DNA-binding -forkhead- domains, enabling spatial reorganization of distant chromosome elements by tethering two DNA molecules together. Yet, domain swapping stability and DNA binding affinity varies between different FoxP proteins. Experimental evidence suggests that the protonation state of a histidine residue conserved in all Fox proteins is responsible for pH-dependent modulation of these interactions. Here, we explore the consequences of the protonation state of another histidine (H59), only conserved within FoxM/O/P subfamilies, on folding and dimerization of the forkhead domain of human FoxP1. Dimer dissociation kinetics and equilibrium unfolding experiments demonstrate that protonation of H59 leads to destabilization of the domain-swapped dimer due to an increase in free energy difference between the monomeric and transition states. This pH-dependence is abolished when H59 is mutated to alanine. Furthermore, anisotropy measurements and molecular dynamics evidence that H59 has a direct impact in the local stability of helix H3. Altogether, our results highlight the relevance of H59 in domain swapping and folding stability of FoxP1.
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PMID:The protonation state of an evolutionarily conserved histidine modulates domainswapping stability of FoxP1. 3297 74