Isotope-edited NMR of cyclosporin A bound to cyclophilin: evidence for a trans 9,10 amide bond.
The binding of a 13C-labeled cyclosporin A (CsA) analog to cyclophilin (peptidyl prolyl isomerase) was examined by means of isotope-edited nuclear magnetic resonance (NMR) techniques. A trans 9,10 peptide bond was adopted when CsA was bound to cyclophilin, in contrast to the cis 9,10 peptide bond found in the crystalline and solution conformations of CsA. Furthermore, nuclear Overhauser effects (NOEs) were observed between the zeta 3 and epsilon 3 protons of the methylleucine (MeLeu) residue at position 9 of CsA and tryptophan121 (Trp121) and phenylalanine (Phe) protons of cyclophilin, suggesting that the MeLeu9 residue of CsA interacts with cyclophilin. These results illustrate the power of isotope-edited NMR techniques for rapidly providing useful information about the conformations and active site environment of inhibitors bound to their target enzymes.
Fesik
S W
SW
Pharmaceutical Discovery Division, Abbott Laboratories, Abbott Park, IL 60064.
Gampe
R T
RT
Holzman
T F
TF
Egan
D A
DA
Edalji
R
R
Luly
J R
JR
Simmer
R
R
Helfrich
R
R
Kishore
V
V
Rich
D H
DH
D000577:Amides; D000591:Amino Acid Isomerases / Q000737:chemistry / Q000378:metabolism*; D002247:Carbon Isotopes; D002352:Carrier Proteins / Q000737:chemistry / Q000378:metabolism*; D003524:Cyclosporins / Q000737:chemistry / Q000378:metabolism*; D004926:Escherichia coli / Q000235:genetics; D006801:Humans; D007930:Leucine / Q000031:analogs & derivatives / Q000737:chemistry; D009682:Magnetic Resonance Spectroscopy / Q000379:methods; D019696:Peptidylprolyl Isomerase; D010649:Phenylalanine / Q000737:chemistry; D011485:Protein Binding; D011487:Protein Conformation; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism; D014364:Tryptophan / Q000737:chemistry
D000577:Amides; D002247:Carbon Isotopes; D002352:Carrier Proteins; D003524:Cyclosporins; D011994:Recombinant Proteins; C030365:leucine methyl ester; D010649:Phenylalanine; D014364:Tryptophan; D000591:Amino Acid Isomerases; D019696:Peptidylprolyl Isomerase; D007930:Leucine