An engineered lipocalin specific for CTLA-4 reveals a combining site with structural and conformational features similar to antibodies

被引:84
作者
Schoenfeld, D. [1 ,2 ]
Matschiner, G. [3 ]
Chatwell, L. [1 ,2 ]
Trentmann, S. [3 ]
Gille, H. [3 ]
Huelsmeyer, M. [3 ]
Brown, N. [4 ,5 ]
Kaye, P. M. [4 ,5 ]
Schlehuber, S. [3 ]
Hohlbaum, A. M. [3 ]
Skerra, A. [1 ,2 ]
机构
[1] Tech Univ Munich, Munich Ctr Integrated Prot Sci, D-85350 Freising Weihenstephan, Germany
[2] Tech Univ Munich, Lehrstuhl Biol Chem, D-85350 Freising Weihenstephan, Germany
[3] Pieris AG, D-85354 Freising Weihenstephan, Germany
[4] Univ York, Ctr Immunol & Infect, Dept Biol, York YO10 5YW, N Yorkshire, England
[5] Univ York, Hull York Med Sch, York YO10 5YW, N Yorkshire, England
关键词
antigen; immunoglobulin; protein crystallography; protein engineering; CYTOTOXIC T-LYMPHOCYTE; CRYSTAL-STRUCTURE; MONOCLONAL-ANTIBODY; LEISHMANIA-DONOVANI; BLOCKADE; ANTIGEN-4; COMPLEX; CANCER; IMMUNOTHERAPY; GLYCOPROTEIN;
D O I
10.1073/pnas.0813399106
中图分类号
O [数理科学和化学]; P [天文学、地球科学]; Q [生物科学]; N [自然科学总论];
学科分类号
07 ; 0710 ; 09 ;
摘要
Biomolecular reagents that enable the specific molecular recognition of proteins play a crucial role in basic research as well as medicine. Up to now, antibodies (immunoglobulins) have been widely used for this purpose. Their predominant feature is the vast repertoire of antigen-binding sites that arise from a set of 6 hypervariable loops. However, antibodies suffer from practical disadvantages because of their complicated architecture, large size, and multiple functions. The lipocalins, on the other hand, have evolved as a protein family that primarily serves for the binding of small molecules. Here, we show that an engineered lipocalin, derived from human Lcn2, can specifically bind the T cell coreceptor CTLA-4 as a prescribed protein target with subnanomolar affinity. Crystallographic analysis reveals that its reshaped cup-like binding site, which is formed by 4 variable loops, provides perfect structural complementarity with this "antigen.'' Furthermore, comparison with the crystal structure of the uncomplexed engineered lipocalin indicates a pronounced induced-fit mechanism, a phenomenon so far considered typical for antibodies. By recognizing the same epitope on CTLA-4 that interacts with the counterreceptors B7.1/B7.2 on antigen-presenting cells the engineered Lcn2 exhibits strong, cross-species antagonistic activity, as evidenced by biological effects comparable with a CTLA-4-specific antibody. With its proven stimulatory activity on T cells in vivo, the CTLA-4 blocking lipocalin offers potential for immunotherapy of cancer and infectious disease. Beyond that, lipocalins with engineered antigen-binding sites, so-called Anticalins, provide a class of small (approximate to 180 residues), structurally simple, and robust binding proteins with applications in the life sciences in general.
引用
收藏
页码:8198 / 8203
页数:6
相关论文
共 33 条
[1]  
[Anonymous], LIPOCALINS
[2]   Small antibody-like proteins with prescribed ligand specificities derived from the lipocalin fold [J].
Beste, G ;
Schmidt, FS ;
Stibora, T ;
Skerra, A .
PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA, 1999, 96 (05) :1898-1903
[3]  
BLAZAR BR, 1994, BLOOD, V83, P3815
[4]   Structural insights into the antigenicity of myelin oligodendrocyte glycoprotein [J].
Breithaupt, C ;
Schubart, A ;
Zander, H ;
Skerra, A ;
Huber, R ;
Linington, C ;
Jacob, U .
PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA, 2003, 100 (16) :9446-9451
[5]   Comparative ligand-binding analysis of ten human lipocalins [J].
Breustedt, DA ;
Schönfeld, DL ;
Skerra, A .
BIOCHIMICA ET BIOPHYSICA ACTA-PROTEINS AND PROTEOMICS, 2006, 1764 (02) :161-173
[6]  
CHEN CY, 1994, J IMMUNOL, V152, P4929
[7]   Crystal structure of a soluble CD28-Fab complex [J].
Evans, EJ ;
Esnouf, RM ;
Manso-Sancho, R ;
Gilbert, RJC ;
James, JR ;
Yu, C ;
Fennelly, JA ;
Vowles, C ;
Hanke, T ;
Walse, B ;
Hünig, T ;
Sorensen, P ;
Stuart, DI ;
Davis, SJ .
NATURE IMMUNOLOGY, 2005, 6 (03) :271-279
[8]   Lipocalin 2 mediates an innate immune response to bacterial infection by sequestrating iron [J].
Flo, TH ;
Smith, KD ;
Sato, S ;
Rodriguez, DJ ;
Holmes, MA ;
Strong, RK ;
Akira, S ;
Aderem, A .
NATURE, 2004, 432 (7019) :917-921
[9]   The neutrophil lipocalin NGAL is a bacteriostatic agent that interferes with siderophore-mediated iron acquisition [J].
Goetz, DH ;
Holmes, MA ;
Borregaard, N ;
Bluhm, ME ;
Raymond, KN ;
Strong, RK .
MOLECULAR CELL, 2002, 10 (05) :1033-1043
[10]   Immunologic and clinical effects of antibody blockade of cytotoxic T lymphocyte-associated antigen 4 in previously vaccinated cancer patients [J].
Hodi, F. Stephen ;
Butler, Marcus ;
Oble, Darryl A. ;
Seiden, Michael V. ;
Haluska, Frank G. ;
Kruse, Andrea ;
MacRae, Suzanne ;
Nelson, Marybeth ;
Canning, Christine ;
Lowy, Israel ;
Korman, Alan ;
Lautz, David ;
Russell, Sara ;
Jaklitsch, Michael T. ;
Ramaiya, Nikhil ;
Chen, Teresa C. ;
Neuberg, Donna ;
Allison, James P. ;
Mihm, Martin C. ;
Dranoff, Glenn .
PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA, 2008, 105 (08) :3005-3010