Crystal structure and biophysical characterization of the nucleoside diphosphate kinase from Leishmania braziliensis

• Grant number: 15/02702-8
• Support Opportunities: Regular Research Grants - Publications - Scientific article
• Start date: May 01, 2015
• End date: October 31, 2015
• Field of knowledge: Biological Sciences - Biochemistry - Chemistry of Macromolecules
• Principal Investigator: Arthur Henrique Cavalcante de Oliveira
• Grantee: Arthur Henrique Cavalcante de Oliveira
• Host Institution: Faculdade de Filosofia, Ciências e Letras de Ribeirão Preto (FFCLRP). Universidade de São Paulo (USP). Ribeirão Preto , SP, Brazil

Abstract

Nucleoside diphosphate kinase (NDK) is a housekeeping enzyme that plays key roles in nucleotide recycling and homeostasis in trypanosomatids. It is also secreted by the intracellular parasite Leishmania to modulate the host response. These functions make NDK an attractive target for drug design and for studies aiming at a better understanding of the mechanisms mediating host-pathogen interactions. We report the crystal structure and biophysical characterization of the NDK from Leishmania braziliensis (LbNDK). The subunit consists of six ±-helices along with a core of four ²-strands arranged in a ²2²3²1²4 antiparallel topology order. In contrast to the NDK from L. major, the LbNDK C-terminal extension is partially unfolded. SAXS data showed that LbNDK forms hexamers in solution in the pH range from 7.0 to 4.0, a hydrodynamic behavior conserved in most eukaryotic NDKs. However, DSF assays show that acidification and alkalization decrease the hexamer stability.Our results support that LbNDK remains hexameric in pH conditions akin to that faced by this enzyme when secreted by Leishmania amastigotes in the parasitophorous vacuoles (pH 4.7 to 5.3). The unusual unfolded conformation of LbNDK C-terminus decreases the surface buried in the trimer interface exposing new regions that might be explored for the development of compounds designed to disturb enzyme oligomerization, which may impair the important nucleotide salvage pathway in these parasites. (AU)