title: Target validation of the inosine monophosphate dehydrogenase (IMPDH) gene in Cryptosporidium using Phylomer(®) peptides.
abstract: Cryptosporidiosis, a gastroenteric disease characterised mainly by diarrheal illnesses in humans and mammals is caused by infection with the protozoan parasite Cryptosporidium. Treatment options for cryptosporidiosis are limited, with the current therapeutic nitazoxanide, only partly efficacious in immunocompetent individuals. The parasite lacks de novo purine synthesis, and is exclusively dependant on purine salvage from its host. Inhibition of the inosine 5' monophosphate dehydrogenase (IMPDH), a purine salvage enzyme that is essential for DNA synthesis, thereby offers a potential drug target against this parasite. In the present study, a yeast-two-hybrid system was used to identify Phylomer peptides within a library constructed from the genomes of 25 phylogenetically diverse bacteria that targeted the IMPDH of Cryptosporidium parvum (IMPcp) and Cryptosporidium hominis (IMPch). We identified 38 unique interacting Phylomers, of which, 12 were synthesised and screened against C. parvum in vitro. Two Phylomers exhibited significant growth inhibition (81.2-83.8% inhibition; P < 0.05), one of which consistently exhibited positive interactions with IMPcp and IMPch during primary and recapitulation yeast two-hybrid screening and did not interact with either of the human IMPDH proteins. The present study highlightsthe potential of Phylomer peptides as target validation tools for Cryptosporidium and other organisms and diseases because of their ability to bind with high affinity to target proteins and disrupt function.
authors.affiliation: School of Veterinary and Life Sciences, Murdoch University, Western Australia, Australia; Phylogica, Telethon Institute for Child Health Research, Subiaco, Western Australia, Australia.
authors.forename: R
authors.initials: R
authors.lastname: Jefferies
authors.affiliation: School of Veterinary and Life Sciences, Murdoch University, Western Australia, Australia.
authors.forename: R
authors.initials: R
authors.lastname: Yang
authors.affiliation: School of Veterinary and Life Sciences, Murdoch University, Western Australia, Australia; Phylogica, Telethon Institute for Child Health Research, Subiaco, Western Australia, Australia.
authors.forename: C K
authors.initials: CK
authors.lastname: Woh
authors.affiliation: School of Veterinary and Life Sciences, Murdoch University, Western Australia, Australia.
authors.forename: T
authors.initials: T
authors.lastname: Weldt
authors.affiliation: Phylogica, Telethon Institute for Child Health Research, Subiaco, Western Australia, Australia.
authors.forename: N
authors.initials: N
authors.lastname: Milech
authors.affiliation: School of Veterinary and Life Sciences, Murdoch University, Western Australia, Australia.
authors.forename: A
authors.initials: A
authors.lastname: Estcourt
authors.affiliation: School of Veterinary and Life Sciences, Murdoch University, Western Australia, Australia.
authors.forename: T
authors.initials: T
authors.lastname: Armstrong
authors.affiliation: Phylogica, Telethon Institute for Child Health Research, Subiaco, Western Australia, Australia.
authors.forename: R
authors.initials: R
authors.lastname: Hopkins
authors.affiliation: Phylogica, Telethon Institute for Child Health Research, Subiaco, Western Australia, Australia.
authors.forename: P
authors.initials: P
authors.lastname: Watt
authors.affiliation: School of Population Health, The University of Queensland, Herston, Queensland, Australia.
authors.forename: S
authors.initials: S
authors.lastname: Reid
authors.affiliation: School of Veterinary and Life Sciences, Murdoch University, Western Australia, Australia.
authors.forename: A
authors.initials: A
authors.lastname: Armson
authors.affiliation: School of Veterinary and Life Sciences, Murdoch University, Western Australia, Australia. Electronic address: una.ryan@murdoch.edu.au.
authors.forename: U M
authors.initials: UM
authors.lastname: Ryan
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D002417:Cattle; D045744:Cell Line, Tumor; D003458:Cryptosporidium / Q000187:drug effects / Q000201:enzymology / Q000235:genetics*; D016054:DNA, Protozoan / Q000302:isolation & purification; D004305:Dose-Response Relationship, Drug; D016680:Genome, Bacterial / Q000235:genetics*; D006801:Humans; D007168:IMP Dehydrogenase / Q000235:genetics* / Q000378:metabolism; D008969:Molecular Sequence Data; D033661:Oocysts; D019151:Peptide Library; D010455:Peptides / Q000737:chemistry / Q000235:genetics* / Q000494:pharmacology / Q000633:toxicity; D010957:Plasmids / Q000235:genetics; D020798:Two-Hybrid System Techniques
chemical_list: D016054:DNA, Protozoan; D019151:Peptide Library; D010455:Peptides; D007168:IMP Dehydrogenase
keywords: Cryptosporidium; Drug target validation; IMPDH; Inosine 5'-monophosphate dehydrogenase; Peptide; Phylomer(®)