title: X-ray structure of snow flea antifreeze protein determined by racemic crystallization of synthetic protein enantiomers.
abstract: Chemical protein synthesis and racemic protein crystallization were used to determine the X-ray structure of the snow flea antifreeze protein (sfAFP). Crystal formation from a racemic solution containing equal amounts of the chemically synthesized proteins d-sfAFP and l-sfAFP occurred much more readily than for l-sfAFP alone. More facile crystal formation also occurred from a quasi-racemic mixture of d-sfAFP and l-Se-sfAFP, a chemical protein analogue that contains an additional -SeCH2- moiety at one residue and thus differs slightly from the true enantiomer. Multiple wavelength anomalous dispersion (MAD) phasing from quasi-racemate crystals was then used to determine the X-ray structure of the sfAFP protein molecule. The resulting model was used to solve by molecular replacement the X-ray structure of l-sfAFP to a resolution of 0.98 A. The l-sfAFP molecule is made up of six antiparallel left-handed PPII helixes, stacked in two sets of three, to form a compact brick-like structure with one hydrophilic face and one hydrophobic face. This is a novel experimental protein structure and closely resembles a structural model proposed for sfAFP. These results illustrate the utility of total chemical synthesis combined with racemic crystallization and X-ray crystallography for determining the unknown structure of a protein.
authors.affiliation: Institute for Biophysical Dynamics, University of Chicago, Chicago, Illinois 60637, USA.
authors.forename: Brad L
authors.initials: BL
authors.lastname: Pentelute
authors.forename: Zachary P
authors.initials: ZP
authors.lastname: Gates
authors.forename: Valentina
authors.initials: V
authors.lastname: Tereshko
authors.forename: Jennifer L
authors.initials: JL
authors.lastname: Dashnau
authors.forename: Jane M
authors.initials: JM
authors.lastname: Vanderkooi
authors.forename: Anthony A
authors.initials: AA
authors.lastname: Kossiakoff
authors.forename: Stephen B H
authors.initials: SB
authors.lastname: Kent
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D021301:Antifreeze Proteins / Q000737:chemistry*; D003460:Crystallization; D018360:Crystallography, X-Ray; D008969:Molecular Sequence Data; D011487:Protein Conformation; D005423:Siphonaptera / Q000737:chemistry*; D013237:Stereoisomerism
chemical_list: D021301:Antifreeze Proteins