Interactions of (AlaAlaLysPro)n and (LysLysSerPro)n with DNA. Proposed coiled-coil structure of AlgR3 and AlgP from Pseudomonas aeruginosa

The proteins, AlgR3 and AlgP, are involved in the regulation of alginate synthesis in Pseudomonas. They contain multiple repeats of Ala*Ala*Lys*Pro as do several other proteins that resemble histones. The interactions of synthesis oligopeptides composed of repeated Ala*Ala*Lys*Pro or Lys*Lys*Ser*Pro units with DNA were studied by fluorescence of the Fmoc (9-fluorenylmethyloxycarbonyl) group attached to the N-termini of the peptides. DNA quenching of the Fmoc fluorescence of the peptides was used to estimate the apparent association constants for the interaction of Fmoc(AAKP)nOH (n = 2, 4, 8, 18, 32) and of Fmoc(KKSP)nOH (n = 2, 4, 8, 16, 20, 32) with DNA. The Fmoc(AAKP)nOH peptides bind to DNA only at low ionic strength; the Fmoc(KKSP)n OH peptides interact with DNA at both low (0.05 M KCl) and high (0.2 M KCl) salt At low ionic strength an increase in the number of the repeat units causes an increase in the apparent association constant up to ˜2 × 106 M−1 for both types of peptides at N ⋍ 24. The insertion of an AAKTA unit into the middle of the Fmoc(AAKP)8OH peptide increases its affinity to DNA. We propose a model of (AAKP)n and of its interaction with DNA. The repeat unit consists of a single turn of α-helix followed by a bend necessitated by Pro. The resultant coiled-coil forms a right-handed superhelix with 10 AAKPs per repeat distance of ˜33 Å. With only slight modification of the canonical parameters of this model the AAKP super helix fits into the major groove of B-form DNA with one AAKP tetramer per base pair repeat of 3.4 Å. The ε-amine nitrogen of Lys can form a polar hydrogen bond with a phosphate oxygen atom of the DNA backbone. A better fit is obtained when the model is modified to accommodate [(AAKP)5AAKTA]n as actually observed in AlgR3. We suggest that this coiled-coil represents a general motif for other protein–DNA interactions.