Binding to human dipeptidyl peptidase IV by adenosine deaminase and antibodies that inhibit ligand binding involves overlapping, discontinuous sites on a predicted beta propeller domain.
Dipeptidyl peptidase IV (DPPIV) is an atypical serine protease that modifies the biological activities of certain chemokines and neuropeptides. In addition, human DPPIV, also known as the T-cell activation antigen CD26, binds adenosine deaminase (ADA) to the T-cell surface, thus protecting the T-cell from adenosine-mediated inhibition of proliferation. Mutations were engineered into DPPIV (five point, 16 single point and six deletion mutations) to examine the binding of ADA and 19 monoclonal antibodies. Deletions of C-terminal residues from the 738-residue extracellular portion of DPPIV showed that the 214 residues C-terminal to Ser552 were not required for ADA binding and that peptidase activity could be ablated by deletion of 20 residues from the C-terminus. Point mutations at either of two locations, Leu294 and Val341, ablated ADA binding. Binding by six anti-DPPIV antibodies that inhibited ADA binding was found to require Leu340 to Arg343 and Thr440/Lys441 but not the 214 residues C-terminal to Ser552. The 13 other antibodies studied bound to a truncated DPPIV consisting of amino acids 1-356. Therefore, the binding sites on DPPIV of ADA and antibodies that inhibit ADA binding are discontinuous and overlapping. Moreover, the 47 and 97 residue spacing of amino acids in these binding sites concords with their location on a beta propeller fold consisting of repeated beta sheets of about 50 amino acids.
Abbott
C A
CA
A.W. Morrow Gastroenterology and Liver Centre, Royal Prince Alfred Hospital, Centenary Institute of Cancer Medicine, University of Sydney, Australia.
McCaughan
G W
GW
Levy
M T
MT
Church
W B
WB
Gorrell
M D
MD
D000243:Adenosine Deaminase / Q000378:metabolism*; D000595:Amino Acid Sequence; D019943:Amino Acid Substitution; D000818:Animals; D000911:Antibodies, Monoclonal; D001665:Binding Sites / Q000235:genetics; D016466:CHO Cells; D006224:Cricetinae; D018819:Dipeptidyl Peptidase 4 / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D000939:Epitopes / Q000737:chemistry / Q000235:genetics; D006801:Humans; D008024:Ligands; D008958:Models, Molecular; D008969:Molecular Sequence Data; D016296:Mutagenesis; D017354:Point Mutation; D017433:Protein Structure, Secondary; D017434:Protein Structure, Tertiary; D051381:Rats; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D017384:Sequence Deletion; D013601:T-Lymphocytes / Q000201:enzymology / Q000276:immunology; D014162:Transfection
D000911:Antibodies, Monoclonal; D000939:Epitopes; D008024:Ligands; D011994:Recombinant Proteins; D018819:Dipeptidyl Peptidase 4; D000243:Adenosine Deaminase