Structure and dynamics of pin1 during catalysis by NMR.
The link between internal enzyme motions and catalysis is poorly understood. Correlated motions in the microsecond-to-millisecond timescale may be critical for enzyme function. We have characterized the backbone dynamics of the peptidylprolyl isomerase (Pin1) catalytic domain in the free state and during catalysis. Pin1 is a prolyl isomerase of the parvulin family and specifically catalyzes the isomerization of phosphorylated Ser/Thr-Pro peptide bonds. Pin1 has been shown to be essential for cell-cycle progression and to interact with the neuronal tau protein inhibiting its aggregation into fibrillar tangles as found in Alzheimer's disease. (15)N relaxation dispersion measurements performed on Pin1 during catalysis reveal conformational exchange processes in the microsecond timescale. A subset of active site residues undergo kinetically similar exchange processes even in the absence of a substrate, suggesting that this area is already "primed" for catalysis. Furthermore, structural data of the turning-over enzyme were obtained through inter- and intramolecular nuclear Overhauser enhancements. This analysis together with a characterization of the substrate concentration dependence of the conformational exchange allowed the distinguishing of regions of the enzyme active site that are affected primarily by substrate binding versus substrate isomerization. Together these data suggest a model for the reaction trajectory of Pin1 catalysis.
Labeikovsky
Wladimir
W
Department of Biochemistry and Howard Hughes Medical Institute, Brandeis University, Waltham, MA 02454, USA.
Eisenmesser
Elan Z
EZ
Bosco
Daryl A
DA
Kern
Dorothee
D
D000595:Amino Acid Sequence; D002384:Catalysis; D015500:Chloramphenicol O-Acetyltransferase / Q000737:chemistry / Q000378:metabolism; D006801:Humans; D007527:Isoenzymes / Q000737:chemistry / Q000378:metabolism; D008954:Models, Biological; D008958:Models, Molecular; D008968:Molecular Conformation; D000072340:NIMA-Interacting Peptidylprolyl Isomerase; D019906:Nuclear Magnetic Resonance, Biomolecular; D019696:Peptidylprolyl Isomerase / Q000737:chemistry* / Q000378:metabolism*; D011485:Protein Binding; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism; D017186:Titrimetry
D007527:Isoenzymes; D000072340:NIMA-Interacting Peptidylprolyl Isomerase; D011994:Recombinant Proteins; D015500:Chloramphenicol O-Acetyltransferase; C000606846:PIN1 protein, human; D019696:Peptidylprolyl Isomerase