Probing the structure and function of human glutaminase-interacting protein: A possible target for drug design

被引:15
作者
Banerjee, Monimoy [2 ]
Huang, Chengdong [2 ]
Marquez, Javier [1 ]
Mohanty, Smita [2 ]
机构
[1] Univ Malaga, Fac Ciencias, Lab Quim Prot, Dept Biol Mol & Bioquim, E-29071 Malaga, Spain
[2] Auburn Univ, Dept Chem & Biochem, Auburn, AL 36849 USA
关键词
D O I
10.1021/bi800287v
中图分类号
Q5 [生物化学]; Q7 [分子生物学];
学科分类号
071010 ; 081704 ;
摘要
PDZ domains are one of the most ubiquitous protein-protein interaction modules found in living systems. Glutaminase interacting protein (GIP), also known as Tax interacting protein 1 (TIP-1), is a PDZ domain-containing protein, which plays pivotal roles in many aspects of cellular signaling, protein scaffolding and modulation of tumor growth. We report here the overexpression, efficient refolding, single-step purification, and biophysical characterization of recombinant human GIP with three different C-terminal target protein recognition sequence motifs by CD, fluorescence, and high-resolution solution NMR methods. It is clear from our NMR analysis that GIP contains 2 alpha-helices and 6 beta-strands. The three target protein C-terminal recognition motifs employed in our interaction studies are glutaminase, beta-catenin and FAS. This is the first report of GIP recognition of the cell surface protein FAS, which belongs to the tumor necrosis factor (TNF) receptor family and mediates cell apoptosis. The dissociation constant (K-D) values for the binding of GIP with different interacting partners as measured by fluorescence spectroscopy range from 1.66 to 2.64 mu M. Significant chemical shift perturbations were observed upon titration of GIP with above three ligands as monitored by 2D (H-1, N-15) -HSQC NMR spectroscopy. GIP undergoes a conformational change upon ligand binding.
引用
收藏
页码:9208 / 9219
页数:12
相关论文
共 52 条
[1]   Overexpression, purification, and characterization of glutaminase-interacting protein, a PDZ-domain protein from human brain [J].
Aledo, JC ;
Rosado, A ;
Olalla, L ;
Campos, JA ;
Márquez, J .
PROTEIN EXPRESSION AND PURIFICATION, 2001, 23 (03) :411-418
[2]   TIP-1 has PDZ scaffold antagonist activity [J].
Alewine, Christine ;
Olsen, Olav ;
Wade, James B. ;
Welling, Paul A. .
MOLECULAR BIOLOGY OF THE CELL, 2006, 17 (10) :4200-4211
[3]   Contribution of single tryptophan residues to the fluorescence and stability of ribonuclease sea [J].
Alston, RW ;
Urbanikova, L ;
Sevcik, J ;
Lasagna, M ;
Reinhart, GD ;
Scholtz, JM ;
Pace, CN .
BIOPHYSICAL JOURNAL, 2004, 87 (06) :4036-4047
[4]   Discs lost, a novel multi-PDZ domain protein, establishes and maintains epithelial polarity [J].
Bhat, MA ;
Izaddoost, S ;
Lu, Y ;
Cho, KO ;
Choi, KW ;
Bellen, HJ .
CELL, 1999, 96 (06) :833-845
[5]  
Cavanagh J., 1996, PROTEIN NMR SPECTROS
[6]  
CHIPMAN DM, 1967, J BIOL CHEM, V242, P4388
[7]   Solution structure and backbone dynamics of the reduced form and an oxidized form of E-coli methionine sulfoxide reductase A (MsrA):: Structural insight of the MsrA catalytic cycle [J].
Coudevylle, Nicolas ;
Antoine, Mathias ;
Bouguet-Bonnet, Sabine ;
Mutzenhardt, Pierre ;
Boschi-Muller, Sandrine ;
Branlant, Guy ;
Cung, Manh-Thong .
JOURNAL OF MOLECULAR BIOLOGY, 2007, 366 (01) :193-206
[8]   NMRPIPE - A MULTIDIMENSIONAL SPECTRAL PROCESSING SYSTEM BASED ON UNIX PIPES [J].
DELAGLIO, F ;
GRZESIEK, S ;
VUISTER, GW ;
ZHU, G ;
PFEIFER, J ;
BAX, A .
JOURNAL OF BIOMOLECULAR NMR, 1995, 6 (03) :277-293
[9]   EXPOSURE OF TRYPTOPHANYL RESIDUES IN PROTEINS - QUANTITATIVE-DETERMINATION BY FLUORESCENCE QUENCHING STUDIES [J].
EFTINK, MR ;
GHIRON, CA .
BIOCHEMISTRY, 1976, 15 (03) :672-680
[10]   SWISS-MODEL and the Swiss-PdbViewer: An environment for comparative protein modeling [J].
Guex, N ;
Peitsch, MC .
ELECTROPHORESIS, 1997, 18 (15) :2714-2723