Talk abstracts

Talk on Saturday 08:50-09:05am submitted by Pakoyo F Kamba

Characterization of an RNA-binding pentatricopeptide repeat protein from Trypanosoma brucei.

Pakoyo F Kamba (Biochemistry and Molecular Biology / Cell and Molecular Biology Program, Michigan State University), Jennifer L Ekstrom (Biochemistry and Molecular Biology, Michigan State University), David A Dickson (Biochemistry and Molecular Biology, Michigan State University), Charles G Hoogstraten (Biochemistry and Molecular Biology / Cell and Molecular Biology Program, Michigan State University)

Abstract:
The flagellate protozoan T. brucei causes an invariably fatal disease, Human African Trypanosomiasis (HAT) or sleeping sickness. Unfortunately, there is no anti-HAT vaccine, yet available drugs are few, highly toxic, and have inadequate efficacy. New, superior drugs are needed, necessitating identification and characterization of novel drug targets. Recently, RNA-binding pentatricopeptide repeat (PPR) proteins were identified to be expanded in number and highly conserved among the trypanosomatids, and hence are potential drug targets. We have embarked on understanding the structure and RNA-binding activity of a 26.9 kDa PPR protein (here called PPR27), the smallest member among at least 28 T. brucei PPR proteins. By refolding of protein from thioredoxin-PPR27 fusion protein inclusion bodies, we were able to determine the regular conformation and oligomeric state of PPR27, as well as obtain preliminary insights into the RNA sequence selectivity of PPR27. We have built on this and expressed soluble PPR27 as a C-terminal fusion to maltose binding protein (MBP). We have generated a panel of N-terminal and C-terminal deletion mutants of PPR27 with the object of elucidating the contribution of the various PPR motifs to RNA binding.

Keywords: T brucei, pentatricopeptide repeat, protein