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MOLECULAR MODELING OF HUMAN GRANULOCYTE-MACROPHAGE COLONY-STIMULATING FACTOR

INTERNATIONAL JOURNAL OF CELL CLONING 1990年第S1期8卷 26-34页

作者： KAUSHANSKY, K BROWN, CB OHARA, PJ ZYMOGENET INC DIV DNA CHEMSEATTLEWA

The colony-stimulating factors (CSFs) are a group of acidic glycoproteins which are required for the proliferation of hematopoietic progenitor cells and for their differentiation into mature blood cells. Receptors for granulocyte-macrophage colony-stimulating factor (GM-CSF) are present on a wide spectrum of cells including erythroid, mixed erythroid-non-erythroid, mixed myeloid and megakaryocytic progenitors, and on mature neutrophils, eosinophils and monocytes. A number of studies are now available which provide insights into the structure-function relationships of human GM-CSF. In an attempt to further understand the interaction between GM-CSF and its cell surface receptor, we have constructed models of the tertiary structure of human GM-CSF using the known disulfide bonding pattern, predictions of the secondary structure of the growth factor and a model based on conformational homologies among cytokines (Parry et al., J Mol Recognition 1988;1:107–110). When compared to a number of functional mapping studies, structural features of the model are consistent with the experimental data, and the model, in turn, leads to the generation of a number of testable hypotheses. The implications of these features in terms of receptor-ligand interaction are discussed.

关键词： Colony-stimulating factors Molecular modeling structure-function relationships Amphiphilicity Helical proteins

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STRUCTURAL HOMOLOGIES BETWEEN LEGUMINOSAE LECTINS AS REVEALED BY THE HYDROPHOBIC CLUSTER-ANALYSIS (HCA) METHOD

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BIOCHEMICAL SYSTEMATICS AND ECOLOGY 1990年第6期18卷 419-427页

作者： ROUGE, P VARLOOT, L UNIV PARIS 06 CNRS MINERAL & CRISTALLOG LAB F-75252 PARIS 05 FRANCE

The Hydrophobic Cluster Analysis (HCA) method, a method that simultaneously allows sequences alignment and secondary structure prediction, was applied to various single- and two-chain Leguminosae lectins. The comparison of the HCA plots shows that all the checked lectins exhibit a very similar dispostion of the strands of .beta.-sheet structure along their amino acid sequence. This is also true for Con A from Canavalia ensiformis and for the Dioclea grandiflora lectin, both from the tribe Diocleae, which are circular permutations of the sequences of other Leguminosae lectins. These results are consistent with the fact that Leguminosae lectins possess a similar overall three-dimensional structure, and indicate that the HCA method yields high levels of information about the conformation of proteins. These results also confirm that Leguminosae lectins constitute a homogeneous group of well-conserved proteins, which may be used as phylogenetic markers for both the precise delineation and the study of the relationship between the lectin-containing Leguminosae taxa.

关键词： Leguminosae lectins structural homologies Hydrophobic Cluster Analysis (HCA) structure-function relationships

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ASPARTATE-70 TO GLYCINE SUBSTITUTION CONFERS RESISTANCE TO NATURALLY-OCCURRING AND SYNTHETIC ANIONIC-SITE LIGANDS ON IN-OVO PRODUCED HUMAN BUTYRYLCHOLINESTERASE

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JOURNAL OF NEUROSCIENCE RESEARCH 1990年第4期27卷 452-460页

作者： NEVILLE, LF GNATT, A LOEWENSTEIN, Y SOREQ, H HEBREW UNIV JERUSALEM INST LIFE SCI DEPT BIOL CHEM IL-91904 JERUSALEM ISRAEL

The "atypical" allelic variant of human butyrylcholinesterase (BuChE) can be characterized by its failure to bind the local anesthetic dibucaine, the muscle relaxant succinylcholine, and the naturally occurring steroidal alkaloid solanidine, all assumed to bind to the charged anionic site component within the normal BuChE enzyme. A single nucleotide substitution conferring a change of aspartate-70 into glycine was recently reported in the CHE gene encoding BuChE from several individuals having the "atypical" BuChE phenotype, whereas in two other DNA samples, this mutation appeared together with a second alteration conferring a change of serine-425 into proline. To separately assess the contribution of each of these mutations toward anionic site interactions in BuChE, three transcription constructs were engineered with each of these substitutions alone or both of them together. Xenopus oocyte microinjection of normal or mutated syntheitc BuChEmRNA transcripts was employed in conjunction with biochemical analyzes of the resultant recombinant BuChE variants. The presence of the Gly-70 mutation alone was found to render the enzyme resistant to 100-mu-M solanidine and 5 mM succinylcholine;concentrations sufficient to inhibit the "normal," Asp-70 containing BuChE by over 50%. Furthermore, when completely inhibited by the organophosphorous poison diisopropylfluorophosphate (DFP), Gly-70 BuChE failed to be reactivated by 10 mM of the cholinesterase-specific oxime pyridine 2-aldoxime methiodide (2-PAM);a concentration restoring about 50% of activity in the "normal" Asp-70 recombinant enzyme. The Pro-425 mutation alone had no apparent influence on BuChE interactions with any of these ligands. However, it conferred synergistic effects on some of the anionic site changes induced by the Gly-70 mutation. Thus, in addition to being resistant to solanidine and to 2-PAM reactivation, the Gly-70/Pro-425 variant completely failed to bind either dibucaine (1 mM) or succinylcholine (1,0

关键词： SERUM CHOLINESTERASE TRANSCRIPTION VECTORS SELECTION ADVANTAGE structure-function relationships INVIVO MUTAGENESIS

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PHYLOGENETICAL ASPECTS OF THE structure-function-relationships OF VERTEBRATE ENKEPHALIN PRECURSORS

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BIOCHEMICAL SYSTEMATICS AND ECOLOGY 1988年第6期16卷 581-587页

作者： ROUGE, P RANFAING, P MAZARGUIL, H CNRS PHARMACOL & TOXICOL FONDAMENTALES LAB F-31400 TOULOUSE FRANCE

Secondary structures and hydropathic profiles of various enkephalin precursors (preproenkephalins A and B, Xenopus proenkephalin), show that the Met- and Leu-enkephalin opioid peptides which they contain are located in buried .beta.-turn-containing regions. The basic amino acid doublets which flank the opioid peptides on both sides are also located in buried regions, which suggests they could act as secondary recognition signals necessary for the proteolytic release of peptides from the precursors. Other basic amino acid sequences located in exposed regions of the precursors could be responsible for the starting proteolytic cleavage of these molecules. This multi-step proteolytic cleavage, depending on the structure of the molecules, appears as a common feature of the well conserved vertebrate enkephalin precursors.

关键词： Enkephalin precursors opioid peptides secondary structure hydropathic profiles proteolytic cleavage structure-function relationships vertebrate protein evolution

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CONFORMATIONAL EFFECTS OF AMINO-ACID SUBSTITUTIONS IN THE P-GLYCOPROTEIN OF THE MDR-1 GENE

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JOURNAL OF PROTEIN CHEMISTRY 1989年第4期8卷 563-573页

作者： BRANDTRAUF, PW LEE, G CARTY, RP PINCUS, MR CHEN, JM SUNY HLTH SCI CTR DEPT PATHOL SYRACUSE NY 13210 USA SUNY DOWNSTATE MED CTR DEPT BIOCHEM BROOKLYN NY 11203 USA COLUMBIA UNIV PRESBYTERIAN MED CTR DEPT MED NEW YORK NY 10032 USA NYU DEPT CHEM NEW YORK NY 10003 USA

The P-glycoprotein of themdr 1 gene is responsible for the phenomenon of multidrug resistance in human cells. The presumed drug-binding site of the wild-type P-glycoprotein contains a glycine at position 185. A mutant P-glycoprotein which contains valine at this position causes cells to retain resistance to colchichine, but to lose cross-resistance to other drugs such as the chemotherapeutic agents vinblastine and Adriamycin. This has been hypothesized to be due to a conformational change in the protein induced by the amino acid substitution. Using conformational energy analysis, we have determined the allowed three-dimensional structures for the wild-type and mutant proteins in the region of position 185. The results indicate that the wild-type protein adopts a unique left-handed conformation at position 185 which is energetically unfavorable for the protein withl-amino acids (including valine) at this position. This conformational change induced by amino acid substitutions for Gly 185 could explain the differences in binding to the P-glycoprotein of various drugs and, hence, the differences in drug resistance exhibited by various cell lines expressing these proteins.

关键词： conformational energy analysis multidrug resistance mutation structure-function relationships three-dimensional structure

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PREDICTIONS OF STRUCTURAL ELEMENTS FOR THE BINDING OF HIN RECOMBINASE WITH THE HIX SITE OF DNA

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PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA 1989年第24期86卷 9841-9845页

作者： PLAXCO, KW MATHIOWETZ, AM GODDARD, WA III Arthur Amos Noyes Laboratory of Chemical Physics California Institute of Technology Pasadena 91125.

Molecular dynamics simulations were coupled with experimental data from biochemistry and genetics to generate a theoretical structure for the binding domain of Hin recombinase complexed with the hix site of DNA. The theoretical model explains the observed sequence specificity of Hin recombinase and leads to a number of testable predictions concerning altered sequence selectivity for various mutants of protein and DNA.

关键词： helix-turn-helix motif molecular modeling Protein-DNA interactions Salmonella typhimurium structure-function relationships

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SUBSTRATE SPECIFICITIES IN CLASS-A BETA-LACTAMASES - PREFERENCE FOR PENAMS VS CEPHEMS - THE ROLE OF RESIDUE-237

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PROTEINS-structure function AND BIOINFORMATICS 1989年第3期6卷 275-283页

作者： HEALEY, WJ LABGOLD, MR RICHARDS, JH CALTECH DIV CHEM & CHEM ENGN 147-75 PASADENA CA 91125 USA

Site saturation mutagenesis has been carried out at Ala-237 in RTEM-1 .beta.-lactamase to assess the role of this site in modulating differences in specificity of .beta.-lactamases for penams vs. cephems as substrates. (An Ala-237 Thr mutation had previously been shown to increase activity on cephems by about 30-80%.1,2) Screening of all 19 possible mutants on penams and cephems revealed the even more active Ala-237 Asn mutant. Detailed kinetic analysis shows that this mutant has about four times the activity toward cephalothin and cephalosporin C as the wild-type enzyme. Both mutations reduce the activity toward penams to about 10% that of RTEM-1 .beta.-lactamase and lower by about ***. C the temperature at which the enzyme denatures. functional properties of the other mutants have also been surveyed. The most interesting aspect of these results is that two quite disparate amino acids, threonine and asparagine, when introduced for Ala-237, cause such similar changes in enzyme specificity while more similar residues do not alter the catalytic properties of the enzyme to such a significant degree.

关键词： mutagenesis structure-function relationships enzymatic catalysis

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structure-function-relationships IN THE PHOTOTOXICITY OF ACETYLENES FROM THE ASTERACEAE

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BIOCHEMICAL SYSTEMATICS AND ECOLOGY 1986年第1期14卷 17-23页

作者： MCLACHLAN, D ARNASON, T LAM, J UNIV OTTAWA DEPT BIOL OTTAWA K1A 6N5 ONTARIO CANADA AARHUS UNIV DEPT ORGAN CHEM DK-8000 AARHUS DENMARK

structure-function relationships of a series of polyacetylenes and thiophene derivatives from the plant family Asteraceae were examined in relation to their photosensitizing activity to Escherichia coli and Saccharomyces cerevisiae cultures. The thiophenes were generally found to be more phototoxic than the polyacetylenes, with increasing activity with increasing number of thiophene rings. With the polyacetylenes there was a general trend of increasing toxicity with increasing acetylene bonds in the molecule. An assessment of the relative phototoxicities of the test compounds revealed a positive correlation between phototoxicity and water-octanol partition coefficients with yeast and to a lesser extent with E. coli. Thiophenes were found to have a higher relative light absorption than acetylenes, but considering all the compounds together there was little correlation between phototoxicity and photon absorption. These results are discussed in relation to modes of action and ecological significance of these phototoxic.

关键词： Asteraceae acetylenes thiophenes phototoxicity structure-function relationships partition coefficient light absorption

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SPECTROSCOPIC AND REDOX PROPERTIES OF PLANT MITOCHONDRIAL CYTOCHROMES-C

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PHYTOCHEMISTRY 1988年第7期27卷 1947-1950页

作者： WALLACE, CJA BOULTER, D UNIV GENEVA DEPT BIOCHIM MED CH-1211 GENEVA 4 SWITZERLAND UNIV DURHAM DEPT BOT DURHAM DH1 3LE ENGLAND

Although the sequences of many higher plant cytochromes c and the crystal structure of the rice protein have been determined, little is known of their functional properties, compared to the wealth of information available for the vertebrate proteins. We have prepared and examined the cytochrome c of three species [Acer negundo, Bixa orellana, Pastinaca sativa], and find that the spectral properties and redox potentials are close to those observed for previously characterized vertebrate and fungal proteins. However, the efficacy of electron transfer by plant cytochromes in reconstituted phosphorylation-coupled terminal oxidation chains from rat mitochondria is markedly lower than that exhibited by either vertebrate or yeast cytochromes c. We have used both similarities and differences to draw conclusions on structure-function relationships in mitochondrial cytochromes c.

关键词： Acer negundo , Bixa orellana , Pastinaca sativa , Chlamydomonas rheinhardtii cytochrome c structure-function relationships spectroscopy redox potential.

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EVALUATION OF THE MUTAGENIC AND CYTOSTATIC POTENTIAL OF ARISTOLOCHIC ACID (3,4-METHYLENEDIOXY-8-METHOXY-10-NITROPHENANTHRENE-1-CARBOXYLIC ACID) AND SEVERAL OF ITS DERIVATIVES

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MUTATION RESEARCH 1988年第4期206卷 447-454页

作者： PEZZUTO, JM SWANSON, SM MAR, W CHE, CT CORDELL, GA FONG, HHS UNIV ILLINOIS COLL PHARM DEPT MED CHEM & PHARMACOGNOSY CHICAGO IL 60612 USA UNIV ILLINOIS COLL MED DIV SURG ONCOL CHICAGO IL 60612 USA

Aristolochic acid (1), a constituent of Aristolochia species, has been used for medicinal purposes since the Graeco-Roman period. Following the observation that the compound was mutagenic and carcinogenic, it was removed from pharmaceutical products. Consistent with previous reports, we have found that 1 serves as a direct-acting mutagen in Salmonella typhimurium strains TA100, TA102, TA1537 and TM677, but was not active in the nitroreductase-deficient strains TA98NR and TA100NR. However, aristolic acid (2), a compound that differs in structure only by the absence of the nitro group, was also found to be a direct-acting mutagen in Salmonella strains TA98, TA100, TA102, TA1537, and TM677, as well as strains TA98NR and TA100NR. Both compounds (1 and 2) were active mutagens when evaluated with cultured Chinese hamster ovary cells. Thus, in contrast to previous suggestions, the nitro group at position 10 is not required to induce a mutagenic response. Also, a series of structural relatives (the methyl esters of 1 and 2 (3 and 4, respectively), aristolochic acid-D (5), aristolactam (6), aristolactam A-II (7), and aristolactam-N-.beta.-D-glucoside (8)) were evaluated for mutagenic potential with Salmonella typhimurium strain TM677 and found to be inactive. Since compounds 3 and 4 were found to be active mutagens with Salmonella typhimurium strains TA98, TA100, TA102 and TA1537 (sufficient quantities of compounds 5-8 were not available for testing), differential sensitivity of the tester strains unrelated to mutagenic potential is suggested. Further, compounds 1, 2, and 6-8 were evaluated for potential to inhibit growth with cultured KB or P388 cells. P388 cells were substantially more sensitive, and compound 1 was the most active of the materials tested (ED5 = 0.58 .mu.M). Compound 6 also demonstrated appreciable activity (ED50 = 4.2 .mu.M), as did compound 8 (ED50 = 6.0 .mu.M). It therefore appears that phenanthrene-ring substituents, in addition to the nitro group at po

关键词： Aristolochic acid Aristolic acid Mutagenicity Salmonella typhimurium nitroreductase-deficient strains structure-function relationships

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