The Cryptosporidium parvum Kinome

J.d Artz ,Wei Qiu,Raymond Hui,Vlad V Lunin,Matthieu Schapira,Yong Zhao,Frank Von Delft,Guillermo Senisterra,Masoud Vedadi,Abdellah Allali-Hassani,Chao Zhang,Irene Chau,A Hutchinson ,Kevan M Shokat ,Yu‐Hui Lin ,P.j Finerty ,Oliver N King ,A.k Wernimont ,M Amani ,J Lew ,A.k Roos ,I Kozieradzki ,F Mackenzie ,Gregory A Wasney ,Tom D Heightman ,O Fedorov

doi:10.1186/1471-2164-12-478

Abstract

BackgroundHundreds of millions of people are infected with cryptosporidiosis annually, with immunocompromised individuals suffering debilitating symptoms and children in socioeconomically challenged regions at risk of repeated infections. There is currently no effective drug available. In order to facilitate the pursuit of anti-cryptosporidiosis targets and compounds, our study spans the classification of the Cryptosporidium parvum kinome and the structural and biochemical characterization of representatives from the CDPK family and a MAP kinase.ResultsThe C. parvum kinome comprises over 70 members, some of which may be promising drug targets. These C. parvum protein kinases include members in the AGC, Atypical, CaMK, CK1, CMGC, and TKL groups; however, almost 35% could only be classified as OPK (other protein kinases). In addition, about 25% of the kinases identified did not have any known orthologues outside of Cryptosporidium spp. Comparison of specific kinases with their Plasmodium falciparum and Toxoplasma gondii orthologues revealed some distinct characteristics within the C. parvum kinome, including potential targets and opportunities for drug design. Structural and biochemical analysis of 4 representatives of the CaMK group and a MAP kinase confirms features that may be exploited in inhibitor design. Indeed, screening CpCDPK1 against a library of kinase inhibitors yielded a set of the pyrazolopyrimidine derivatives (PP1-derivatives) with IC50 values of < 10 nM. The binding of a PP1-derivative is further described by an inhibitor-bound crystal structure of CpCDPK1. In addition, structural analysis of CpCDPK4 identified an unprecedented Zn-finger within the CDPK kinase domain that may have implications for its regulation.ConclusionsIdentification and comparison of the C. parvum protein kinases against other parasitic kinases shows how orthologue- and family-based research can be used to facilitate characterization of promising drug targets and the search for new drugs.

Highlights

Hundreds of millions of people are infected with cryptosporidiosis annually, with immunocompromised individuals suffering debilitating symptoms and children in socioeconomically challenged regions at risk of repeated infections
Breakdown of the Cryptosporidium parvum kinome Assignment of the protein kinases to their subfamilies was accomplished via clustering of the kinase domain by sequence similarity
We found 73 protein kinases with intact catalytic triads [20], including those falling into the following categories: AGC, CaMK, CK1, CMGC, TKL, Atypical, and OPK

Summary

Introduction

Hundreds of millions of people are infected with cryptosporidiosis annually, with immunocompromised individuals suffering debilitating symptoms and children in socioeconomically challenged regions at risk of repeated infections. C. parvum is an obligate parasite in the same phylum of Apicomplexa as Plasmodium and the same order of Eucoccidiorida as Toxoplasma and Eimeria It is one of the pathogenic agents responsible of cryptosporidiosis as a neglected disease, one that lacks an effective drug [6]. To the best of our knowledge, there have been only 4 studies to date in which a Cryptosporidium target and effective inhibitors have been identified and characterized These include inosine 5’-monophosphate dehydrogenase [10], S-adenosylhomocysteine hydrolase [11], nonspecific polyprenyl pyrophosphate synthase (related to farnesyl pyrophosphate synthase) [12] and calciumdependent protein kinase 1 (CDPK1) [13], where the latter two targets were contributed by structural genomics groups. Comparison of the CpPKs (protein kinases) with other known parasitic kinases illustrates some of their unique features and demonstrates that there are potential drug targets, as well as opportunities for drug design

Methods

Results

Conclusion