Heparan Sulfate and Heparin Promote Faithful Prion Replication in Vitro by Binding to Normal and Abnormal Prion Proteins in Protein Misfolding Cyclic Amplification.
The precise mechanism underlying the conversion of normal prion protein (PrPC) into abnormal prion protein (PrPSc) remains unclear. Protein misfolding cyclic amplification (PMCA), an in vitro technique used for amplifying PrPSc, results in PrPScreplication that preserves the strain-specific characteristics of the input PrPSc; thus, PMCA mimics the process of in vivo PrPScreplication. Previous work has demonstrated that in PMCA, nucleic acids are critical for PrPScamplification, but little information has been reported on glycosaminoglycan (GAG) participation in PrPScreplication in vitro Here, we investigated whether GAGs play a role in the faithful replication of PrPScby using a modified PMCA performed with baculovirus-derived recombinant PrP (Bac-PrP) as a substrate. The addition of heparan sulfate (HS) or its analog heparin (HP) restored the conversion efficiency in PMCA that was inhibited through nucleic acid depletion. Moreover, the PMCA products obtained under these conditions were infectious and preserved the properties of the input PrPScThese data suggest that HS and HP play the same role as nucleic acids in facilitating faithful replication of prions in PMCA. Furthermore, we showed that HP binds to both Bac-PrP and Bac-PrPScthrough the sulfated groups present on HP and that the N-terminal domain of Bac-PrPScmight potentially not be involved in the binding to HP. These results suggest that the interaction of GAGs such as HS and HP with PrPCand/or PrPScthrough their sulfate groups is critical for the faithful replication of prions.
From the National Institute of Animal Health, National Agriculture and Food Research Organization (NARO), Tsukuba, Ibaraki 305-0856, Japan imamuram@affrc.go.jp.
Morikazu
M
Imamura
From the National Institute of Animal Health, National Agriculture and Food Research Organization (NARO), Tsukuba, Ibaraki 305-0856, Japan.
Naoko
N
Tabeta
From the National Institute of Animal Health, National Agriculture and Food Research Organization (NARO), Tsukuba, Ibaraki 305-0856, Japan.
Nobuko
N
Kato
From the National Institute of Animal Health, National Agriculture and Food Research Organization (NARO), Tsukuba, Ibaraki 305-0856, Japan.
Yuichi
Y
Matsuura
From the National Institute of Animal Health, National Agriculture and Food Research Organization (NARO), Tsukuba, Ibaraki 305-0856, Japan.
Yoshifumi
Y
Iwamaru
From the National Institute of Animal Health, National Agriculture and Food Research Organization (NARO), Tsukuba, Ibaraki 305-0856, Japan.
Takashi
T
Yokoyama
From the National Institute of Animal Health, National Agriculture and Food Research Organization (NARO), Tsukuba, Ibaraki 305-0856, Japan.
Yuichi
Y
Murayama
D000818:Animals; D006493:Heparin / Q000737:chemistry*; D006497:Heparan Sulfate / Q000737:chemistry*; D051379:Mice; D000072002:Prion Proteins / Q000737:chemistry*; D000072417:Protein Domains; D017510:Protein Folding
D006493:Heparin; D006497:Heparan Sulfate; D000072002:Prion Proteins
baculovirus-recombinant prion protein; heparan sulfate; heparin; prion; prion disease; protein folding; protein misfolding cyclic amplification