Extended binding sites of cyclophilin as revealed by the interaction with HIV‐1 Gag polyprotein derived oligopeptides

FEBS Letters

Reimer, Ulf¹  Schutkowski, Mike¹  Jakob, Mario¹  Wöllner, Steffen¹  Scherer, Gerd¹  Drewello, Mario¹  Schierhorn, Angelika¹  Fischer, Gunter¹ 

[1] Max-Planck-Gesellschaft zur Förderung der Wissenschaften e. V., Arbeits gruppe ‘Enzymologie der Peptidbindung’ Kurt-Mothes-Straβe 3, D-06120 Halle/Saale, Germany

关键词: Cyclophilin;    HIV-1 Gag polyprotein;    Peptide;    CD spectroscopy;    Selective binding;    Cyp18;    human recombinant cytosolic cyclophilin with molecular mass of 18 kDa (according to FIscher;    G. (1994) Angew. Chem. Int. Ed. Engl. 33;    1415–1436);    PPIase;    peptidyl prolyl cisltrans isomerase;    FKBP;    FK506 binding protein;    CsA;    cyclosporin A;    HIV-1;    human immunodeficiency virus type 1;    SIV;    simian immunodeficiency virus;    NMR;    nuclear magnetic resonance;    TFA;    trifluoroacetic acid;    DCM;    dichloromethane;    DIEA;    N;    N-diisopropylethylamine;    DMAP;    N;    N-dimethylaminopyridine;    Fmoc;    9-fluorenylmethoxycarbonyl;    GdmCl;    guanidinium hydrochloride;    HBTU;    O-(benzotriazol1-yl)-1;    1;    3;    3-tetramethyluronium hexafluorophosphate;    HATU;    O-(7-azabenzotriazol-1-yl)-1;    1;    3;    3-tetramethyluronium hexafluorophosphate;    HOAt;    1-hydroxy-7-azabenzotriazole;    HPLC;    high performance liquid chromatography;    MBHA;    4-methylbenzhydrylamine;    NMP;    N-methylpyrrolidone;    tBu;    tert.-butyl;    Trt;    trityl;    HOBt;    1hydroxybenzotriazole;    Pmc;    2;    2;    5;    7;    8-pentamethylchroman-6-sulfonyl;    Ac;    acetyl;    CE;    capillary electrophoresis;    CA;    capsid protein of HIV-1;    GST;    glutathione S-transferase;   

DOI  :  10.1016/0014-5793(96)00972-6

学科分类：生物化学/生物物理

来源: John Wiley & Sons Ltd.

【 摘 要 】

Oligopeptides derived from the gag polyprotein (Pr55^gag) of human immunodeficiency virus type 1 (HIV-1) segment were used to evaluate the extension of the putative binding region for the complex of Pr55^gag and the human cytosolic peptidyl prolyl cisltrans isomerase (PPIase) 18 kDa cyclophilin (Cyp18). Five N-terminally acetylated, C-terminally amidated oligopeptides containing one (HIV-1 Gag ^218–224; 1), two (HIV-1 Gag ^218–226 and HIV-1 Gag ^217−224; 2 and 3, respectively), three (HIV-1 Gag ^217–226; 4) or four (HIV-1 Gag^213−237; 5) proline residues were synthesized. Using competition experiments with a standard substrate the binding affinities to Cyp18 of the synthesized peptides were determined. The IC₅₀ value of 184 μ.M for the 25-mer peptide 5 was fivefold or more lower than those of the peptides 1–4 lacking one or more prolines. Failure of competition in assays containing enzymes of other PPIase families by millimolar concentrations of 5 revealed a Cyp18 specific interaction involving the active site of the enzyme. In its far UV circular dichroism, aqueous solutions of 5 display properties of random coil conformation, but spectra were also consistent with a small contribution of proline specific secondary structures. However, a proline-rich peptide typical of forming left-handed polyproline II helices did not compete for the active site of Cyp18. The results demonstrate that the putative binding region of HIV-1 gag polyprotein has a certain degree of binding affinity to the PPIase site of Cyp18, and may add a previously unrecognized topological component to the known subsite specificity of cyclophilins.

【 授权许可】

Unknown