Molecular basis for an ancient partnership between prolyl isomerase Pin1 and phosphatase inhibitor-2

Biochemistry. 2011 Aug 2;50(30):6567-78. doi: 10.1021/bi200553e. Epub 2011 Jul 8.


Pin1 is a prolyl isomerase that recognizes phosphorylated Ser/Thr-Pro sites, and phosphatase inhibitor-2 (I-2) is phosphorylated during mitosis at a PSpTP site that is expected to be a Pin1 substrate. However, we previously discovered I-2, but not phospho-I-2, bound to Pin1 as an allosteric modifier of Pin1 substrate specificity [Li, M., et al. (2008) Biochemistry 47, 292]. Here, we use binding assays and NMR spectroscopy to map the interactions on Pin1 and I-2 to elucidate the organization of this complex. Despite having sequences that are ∼50% identical, human, Xenopus, and Drosophila I-2 proteins all exhibited identical, saturable binding to GST-Pin1 with K(0.5) values of 0.3 μM. The (1)H-(15)N heteronuclear single-quantum coherence spectra for both the WW domain and isomerase domain of Pin1 showed distinctive shifts upon addition of I-2. Conversely, as shown by NMR spectroscopy, specific regions of I-2 were affected by addition of Pin1. A single-residue I68A substitution in I-2 weakened binding to Pin1 by half and essentially eliminated binding to the isolated WW domain. On the other hand, truncation of I-2 to residue 152 had a minimal effect on binding to the WW domain but eliminated binding to the isomerase domain. Size exclusion chromatography revealed that wild-type I-2 and Pin1 formed a large (>300 kDa) complex and I-2(I68A) formed a complex of half the size that we propose are a heterotetramer and a heterodimer, respectively. Pin1 and I-2 are conserved among eukaryotes from yeast to humans, and we propose they make up an ancient partnership that provides a means for regulating Pin1 specificity and function.

Publication types

  • Comparative Study
  • Research Support, N.I.H., Extramural

MeSH terms

  • Allosteric Regulation / genetics
  • Amino Acid Sequence
  • Animals
  • Conserved Sequence
  • Drosophila Proteins / chemistry*
  • Drosophila Proteins / metabolism
  • Escherichia coli Proteins / chemistry
  • Escherichia coli Proteins / genetics
  • Humans
  • Molecular Sequence Data
  • NIMA-Interacting Peptidylprolyl Isomerase
  • Peptidylprolyl Isomerase / chemistry*
  • Peptidylprolyl Isomerase / metabolism
  • Phosphoprotein Phosphatases / antagonists & inhibitors*
  • Phosphoprotein Phosphatases / chemistry
  • Phosphoprotein Phosphatases / metabolism
  • Phosphorylation
  • Protein Binding
  • Protein Multimerization
  • Proteins / chemistry
  • Proteins / metabolism*
  • Saccharomyces cerevisiae / enzymology
  • Saccharomyces cerevisiae Proteins
  • Sequence Homology, Amino Acid*
  • Substrate Specificity / genetics
  • Xenopus Proteins / chemistry*
  • Xenopus Proteins / metabolism


  • Drosophila Proteins
  • Escherichia coli Proteins
  • NIMA-Interacting Peptidylprolyl Isomerase
  • Proteins
  • Saccharomyces cerevisiae Proteins
  • Xenopus Proteins
  • protein phosphatase inhibitor-2
  • Phosphoprotein Phosphatases
  • ESS1 protein, S cerevisiae
  • PIN1 protein, human
  • Peptidylprolyl Isomerase
  • dod protein, Drosophila
  • parvA protein, E coli
  • Pin1 protein, Xenopus