title: Biochemical Analysis of Interaction between Kringle Domains of Plasminogen and Prion Proteins with Q167R Mutation.
abstract: The conformational change of cellular prion protein (PrPC) to its misfolded counterpart, termed PrPSc, is mediated by a hypothesized cellular cofactor. This cofactor is believed to interact directly with certain amino acid residues of PrPC. When these are mutated into cationic amino acid residues, PrPScformation and prion replication halt in a dominant negative (DN) manner, presumably due to strong binding of the cofactor to mutated PrPC, designated as DN PrP mutants. Previous studies demonstrated that plasminogen and its kringle domains bind to PrP and accelerate PrPScgeneration. In this study, in vitro binding analysis of kringle domains of plasminogen to Q167R DN mutant PrP (PrPQ167R) was performed in parallel with the wild type (WT) and Q218K DN mutant PrP (PrPQ218K). The binding affinity of PrPQ167R was higher than that of WT PrP, but lower than that of PrPQ218K. Scatchard analysis further indicated that, like PrPQ218K and WT PrP, PrPQ167R interaction with plasminogen occurred at multiple sites, suggesting cooperativity in this interaction. Competitive binding analysis usingL-lysine orL-arginine confirmed the increase of the specificity and binding affinity of the interaction as PrP acquired DN mutations. Circular dichroism spectroscopy demonstrated that the recombinant PrPs used in this study retained the α-helix-rich structure. The α-helix unfolding study revealed similar conformational stability for WT and DN-mutated PrPs. This study provides an additional piece of biochemical evidence concerning the interaction of plasminogen with DN mutant PrPs.
authors.affiliation: Division of Zoonoses, Center for Immunology and Pathology, National Institute of Health, Korea Centers for Disease Control and Prevention, Cheongwon 28159, Republic of Korea.
authors.forename: Jeongmin
authors.initials: J
authors.lastname: Lee
authors.affiliation: Department of Genetic Engineering, Sungkyunkwan University, Suwon 16419, Republic of Korea.
authors.forename: Byoung Woo
authors.initials: BW
authors.lastname: Lee
authors.affiliation: Division of Foreign Animal Disease, Animal and Plant Quarantine Agency, Gimcheon 39660, Republic of Korea.
authors.forename: Hae-Eun
authors.initials: HE
authors.lastname: Kang
authors.affiliation: Department of Pharmacy and Institute of Pharmaceutical Science and Technology, Hanyang University, Ansan 15588, Republic of Korea.
authors.forename: Kevine K
authors.initials: KK
authors.lastname: Choe
authors.affiliation: Department of Genetic Engineering, Sungkyunkwan University, Suwon 16419, Republic of Korea.
authors.forename: Moosik
authors.initials: M
authors.lastname: Kwon
authors.affiliation: Department of Pharmacy and Institute of Pharmaceutical Science and Technology, Hanyang University, Ansan 15588, Republic of Korea.
authors.forename: Chongsuk
authors.initials: C
authors.lastname: Ryou
mesh_terms: D000596:Amino Acids / Q000032:analysis; D000818:Animals; D001419:Bacteria / Q000235:genetics; D001483:Base Sequence; D003001:Cloning, Molecular; D003067:Coenzymes / Q000378:metabolism; D004926:Escherichia coli / Q000235:genetics; D005786:Gene Expression Regulation; D018082:Kringles; D051379:Mice; D016297:Mutagenesis, Site-Directed; D009154:Mutation; D010958:Plasminogen / Q000378:metabolism*; D000072002:Prion Proteins / Q000235:genetics* / Q000378:metabolism*; D011485:Protein Binding; D011487:Protein Conformation; D017510:Protein Folding; D054730:Protein Interaction Domains and Motifs; D011994:Recombinant Proteins
chemical_list: D000596:Amino Acids; D003067:Coenzymes; D000072002:Prion Proteins; D011994:Recombinant Proteins; D010958:Plasminogen
keywords: cofactor; dominant negative mutant; plasminogen; prion protein