title: Mono-anionic phosphopeptides produced by unexpected histidine alkylation exhibit high Plk1 polo-box domain-binding affinities and enhanced antiproliferative effects in HeLa cells.
abstract: Binding of polo-like kinase 1 (Plk1) polo-box domains (PBDs) to phosphothreonine (pThr)/phosphoserine (pSer)-containing sequences is critical for the proper function of Plk1. Although high-affinity synthetic pThr-containing peptides provide starting points for developing PBD-directed inhibitors, to date the efficacy of such peptides in whole cell assays has been poor. This potentially reflects limited cell membrane permeability arising, in part, from the di-anionic nature of the phosphoryl group or its mimetics. In our current article we report the unanticipated on-resin N(τ)-alkylation of histidine residues already bearing a N(π)- alkyl group. This resulted in cationic imidazolium-containing pThr peptides, several of which exhibit single-digit nanomolar PBD-binding affinities in extracellular assays and improved antimitotic efficacies in intact cells. We enhanced the cellular efficacies of these peptides further by applying bio-reversible pivaloyloxymethyl (POM) phosphoryl protection. New structural insights presented in our current study, including the potential utility of intramolecular charge masking, may be useful for the further development of PBD-binding peptides and peptide mimetics.
authors.affiliation: Chemical Biology Laboratory, Center for Cancer Research, National Institutes of Health, NCI at Frederick, Frederick, MD, 21702.
authors.forename: Wen-Jian
authors.initials: WJ
authors.lastname: Qian
authors.forename: Jung-Eun
authors.initials: JE
authors.lastname: Park
authors.forename: Dan
authors.initials: D
authors.lastname: Lim
authors.forename: Christopher C
authors.initials: CC
authors.lastname: Lai
authors.forename: James A
authors.initials: JA
authors.lastname: Kelley
authors.forename: Suk-Youl
authors.initials: SY
authors.lastname: Park
authors.forename: Ki Won
authors.initials: KW
authors.lastname: Lee
authors.forename: Michael B
authors.initials: MB
authors.lastname: Yaffe
authors.forename: Kyung S
authors.initials: KS
authors.lastname: Lee
authors.forename: Terrence R
authors.initials: TR
authors.lastname: Burke
mesh_terms: D000478:Alkylation; D000838:Anions; D018797:Cell Cycle Proteins / Q000737:chemistry* / Q000378:metabolism*; D049109:Cell Proliferation / Q000187:drug effects; D002853:Chromatography, Liquid; D003460:Crystallization; D004795:Enzyme Stability / Q000187:drug effects; D004797:Enzyme-Linked Immunosorbent Assay; D004950:Esterases / Q000378:metabolism; D005454:Fluorescence Polarization; D006367:HeLa Cells; D006639:Histidine / Q000737:chemistry / Q000378:metabolism*; D006801:Humans; D010748:Phosphopeptides / Q000138:chemical synthesis* / Q000737:chemistry / Q000494:pharmacology*; D011355:Prodrugs / Q000494:pharmacology; D011485:Protein Binding / Q000187:drug effects; D017346:Protein Serine-Threonine Kinases / Q000737:chemistry* / Q000378:metabolism*; D017434:Protein Structure, Tertiary; D011518:Proto-Oncogene Proteins / Q000737:chemistry* / Q000378:metabolism*; D053719:Tandem Mass Spectrometry; D000097869:Polo-Like Kinase 1
chemical_list: D000838:Anions; D018797:Cell Cycle Proteins; D004950:Esterases; D006639:Histidine; D010748:Phosphopeptides; D011355:Prodrugs; D017346:Protein Serine-Threonine Kinases; D011518:Proto-Oncogene Proteins; D000097869:Polo-Like Kinase 1
keywords: Plk1; cationic dialkyl histidine; crystal structure; intramolecular charge masking; phosphopeptides; polo kinase; polo-box domain