The role of the C-terminus and Kpn loop in the quaternary structure stability of nucleoside diphosphate kinase from Leishmania parasites

Author(s)

Vieira, Plinio Salmazo ^[1] ; de Giuseppe, Priscila Oliveira ^[1] ; Cavalcante de Oliveira, Arthur Henrique ^[2] ; Murakami, Mario Tyago ^[1]

Affiliation

^[1] CNPEM, Lab Nacl Biociencias, Campinas, SP - Brazil

^[2] Univ Sao Paulo, Fac Filosofia Ciencias & Letras Ribeirao Preto, Dept Quim, BR-14049 Ribeirao Preto - Brazil

• Document type: Journal article
• Source: Journal of Structural Biology; v. 192, n. 3, p. 336-341, DEC 2015.
• Web of Science Citations: 2

Abstract

Nucleoside diphosphate kinase (NDK) is a housekeeping enzyme that plays key roles in nucleotide recycling and homeostasis in trypanosomatids. Moreover, it is 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. Here, we report the crystal structures of three mutants of the NDK from Leishmania major (LmNDK) that affects the stability of the hexameric biological assembly including P95S, Delta 5Ct (lacking the last five residues) and the double mutant P100S/Delta 5Ct. Although P95S and Delta 5Ct variants conserve the hexameric structure of the wild-type protein, the double mutant becomes a dimer as shown by in solution studies. Free energy calculation of dimer-dimer interfaces and enzymatic assays indicate that P95S, Delta 5Ct and P100S/Delta 5Ct mutations progressively decrease the hexamer stability and enzyme activity. These results demonstrate that the mutated regions play a role in protein function through stabilizing the quaternary arrangement. (C) 2015 Elsevier Inc. All rights reserved. (AU)

• FAPESP's process: 11/20569-2 - Role of amino acids of the active site and interface in oligomeric in the structure and function of the nucleoside diphosphate kinase of Leishmania major
• Grantee: Arthur Henrique Cavalcante de Oliveira
• Support Opportunities: Regular Research Grants
• FAPESP's process: 10/03761-4 - Kinetic characterization and effect of mutations in the oligomeric interface of the Nucleoside Diphosphate Kinase from Leishmania major.
• Grantee: Plínio Salmazo Vieira
• Support Opportunities: Scholarships in Brazil - Master
• FAPESP's process: 07/06755-2 - Structural and functional characterization of proteins envolved in the virulence of the protozoan parasite Leishmania major
• Grantee: Arthur Henrique Cavalcante de Oliveira
• Support Opportunities: Regular Research Grants
• FAPESP's process: 11/24178-8 - Structural studies of NEK kinases from Leishmania braziliensis.
• Grantee: Plínio Salmazo Vieira
• Support Opportunities: Scholarships in Brazil - Doctorate
• FAPESP's process: 10/51730-0 - Functional and structural studies of protein kinases involved in cancer and neglected diseases: towards the development of new inhibitors
• Grantee: Jörg Kobarg
• Support Opportunities: Regular Research Grants