title: Molecular basis for the broad substrate selectivity of a peptide prenyltransferase.
abstract: The cyanobactin prenyltransferases catalyze a series of known or unprecedented reactions on millions of different substrates, with no easily observable recognition motif and exquisite regioselectivity. Here we define the basis of broad substrate tolerance for the otherwise uncharacterized TruF family. We determined the structures of the Tyr-prenylating enzyme PagF, in complex with an isoprenoid donor analog and a panel of linear and macrocyclic peptide substrates. Unexpectedly, the structures reveal a truncated barrel fold, wherein binding of large peptide substrates is necessary to complete a solvent-exposed hydrophobic pocket to form the catalytically competent active site. Kinetic, mutational, chemical, and computational analyses revealed the structural basis of selectivity, showing a small motif within peptide substrates that is sufficient for recognition by the enzyme. Attaching this 2-residue motif to two random peptides results in their isoprenylation by PagF, demonstrating utility as a general biocatalytic platform for modifications on any peptide substrate.
authors.affiliation: Department of Biochemistry, University of Illinois at Urbana-Champaign, Urbana, IL 61801.
authors.forename: Yue
authors.initials: Y
authors.lastname: Hao
authors.affiliation: Department of Medicinal Chemistry, University of Utah, Salt Lake City, UT 84112.
authors.forename: Elizabeth
authors.initials: E
authors.lastname: Pierce
authors.affiliation: Department of Medicinal Chemistry, University of Utah, Salt Lake City, UT 84112.
authors.forename: Daniel
authors.initials: D
authors.lastname: Roe
authors.affiliation: Department of Medicinal Chemistry, University of Utah, Salt Lake City, UT 84112.
authors.forename: Maho
authors.initials: M
authors.lastname: Morita
authors.affiliation: Department of Medicinal Chemistry, University of Utah, Salt Lake City, UT 84112.
authors.forename: John A
authors.initials: JA
authors.lastname: McIntosh
authors.affiliation: Department of Biochemistry, University of Illinois at Urbana-Champaign, Urbana, IL 61801.|Center for Biophysics and Computational Biology, University of Illinois at Urbana-Champaign, Urbana, IL 61801.
authors.forename: Vinayak
authors.initials: V
authors.lastname: Agarwal
authors.affiliation: Department of Medicinal Chemistry, University of Utah, Salt Lake City, UT 84112.
authors.forename: Thomas E
authors.initials: TE
authors.lastname: Cheatham
authors.affiliation: Department of Medicinal Chemistry, University of Utah, Salt Lake City, UT 84112; ews1@utah.edu snair@illinois.edu.
authors.forename: Eric W
authors.initials: EW
authors.lastname: Schmidt
authors.affiliation: Department of Biochemistry, University of Illinois at Urbana-Champaign, Urbana, IL 61801; ews1@utah.edu snair@illinois.edu.|Center for Biophysics and Computational Biology, University of Illinois at Urbana-Champaign, Urbana, IL 61801.|Institute for Genomic Biology, University of Illinois at Urbana-Champaign, Urbana, IL 61801.
authors.forename: Satish K
authors.initials: SK
authors.lastname: Nair
mesh_terms: D020816:Amino Acid Motifs; D000595:Amino Acid Sequence; D020134:Catalytic Domain; D018360:Crystallography, X-Ray; D004122:Dimethylallyltranstransferase / Q000235:genetics / Q000378:metabolism*; D010455:Peptides / Q000737:chemistry; D054876:Prenylation; D011485:Protein Binding; D013329:Structure-Activity Relationship; D013379:Substrate Specificity
chemical_list: D010455:Peptides; D004122:Dimethylallyltranstransferase
keywords: RiPP; biosynthesis; crystallography; prenylation