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Solution structures of polcalcin Phl p 7 in three ligation states: Apo‐, hemi‐Mg 2+ ‐bound, and fully Ca 2+ ‐bound
Author(s) -
Henzl Michael T.,
Sirianni Arthur G.,
Wycoff Wei G.,
Tan Anmin,
Tanner John J.
Publication year - 2013
Publication title -
proteins: structure, function, and bioinformatics
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.699
H-Index - 191
eISSN - 1097-0134
pISSN - 0887-3585
DOI - 10.1002/prot.24186
Subject(s) - chemistry , dimer , carboxylate , crystallography , stereochemistry , ligand (biochemistry) , antiparallel (mathematics) , divalent , crystal structure , monomer , protein structure , biochemistry , receptor , physics , organic chemistry , quantum mechanics , magnetic field , polymer
Polcalcins are small EF‐hand proteins believed to assist in regulating pollen‐tube growth. Phl p 7, from timothy grass ( Phleum pratense ), crystallizes as a domain‐swapped dimer at low pH. This study describes the solution structures of the recombinant protein in buffered saline at pH 6.0, containing either 5.0 m M EDTA, 5.0 m M Mg 2+ , or 100 μ M Ca 2+ . Phl p 7 is monomeric in all three ligation states. In the apo‐form, both EF‐hand motifs reside in the closed conformation, with roughly antiparallel N‐ and C‐terminal helical segments. In 5.0 m M Mg 2+ , the divalent ion is bound by EF‐hand 2, perturbing interhelical angles and imposing more regular helical structure. The structure of Ca 2+ ‐bound Phl p 7 resembles that previously reported for Bet v 4—likewise exposing apolar surface to the solvent. Occluded in the apo‐ and Mg 2+ ‐bound forms, this surface presumably provides the docking site for Phl p 7 targets. Unlike Bet v 4, EF‐hand 2 in Phl p 7 includes five potential anionic ligands, due to replacement of the consensus serine residue at – x (residue 55 in Phl p 7) with aspartate. In the Phl p 7 crystal structure, D55 functions as a helix cap for helix D. In solution, however, D55 apparently serves as a ligand to the bound Ca 2+ . When Mg 2+ resides in site 2, the D55 carboxylate withdraws to a distance consistent with a role as an outer‐sphere ligand. 15 N relaxation data, collected at 600 MHz, indicate that backbone mobility is limited in all three ligation states. Proteins 2013. © 2012 Wiley Periodicals, Inc.