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Structures of aspartate aminotransferases from Trypanosoma brucei, Leishmania major and Giardia lamblia.

ABSTRACT: The structures of three aspartate aminotransferases (AATs) from eukaryotic pathogens were solved within the Seattle Structural Genomics Center for Infectious Disease (SSGCID). Both the open and closed conformations of AAT were observed. Pyridoxal phosphate was bound to the active site via a Schiff base to a conserved lysine. An active-site mutant showed that Trypanosoma brucei AAT still binds pyridoxal phosphate even in the absence of the tethering lysine. The structures highlight the challenges for the structure-based design of inhibitors targeting the active site, while showing options for inhibitor design targeting the N-terminal arm.

SUBMITTER: Abendroth J

PROVIDER: S-EPMC4427166 | biostudies-literature | 2015 May

REPOSITORIES: biostudies-literature

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Structures of aspartate aminotransferases from Trypanosoma brucei, Leishmania major and Giardia lamblia.

Abendroth Jan J Choi Ryan R Wall Abigail A Clifton Matthew C MC Lukacs Christine M CM Staker Bart L BL Van Voorhis Wesley W Myler Peter P Lorimer Don D DD Edwards Thomas E TE

Acta crystallographica. Section F, Structural biology communications 20150421 Pt 5

The structures of three aspartate aminotransferases (AATs) from eukaryotic pathogens were solved within the Seattle Structural Genomics Center for Infectious Disease (SSGCID). Both the open and closed conformations of AAT were observed. Pyridoxal phosphate was bound to the active site via a Schiff base to a conserved lysine. An active-site mutant showed that Trypanosoma brucei AAT still binds pyridoxal phosphate even in the absence of the tethering lysine. The structures highlight the challenges ...[more]

PMID: 25945710

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Project description:BackgroundThe trypanosomatids Leishmania major, Trypanosoma brucei and Trypanosoma cruzi cause some of the most debilitating diseases of humankind: cutaneous leishmaniasis, African sleeping sickness, and Chagas disease. These protozoa possess complex life cycles that involve development in mammalian and insect hosts, and a tightly coordinated cell cycle ensures propagation of the highly polarized cells. However, the ways in which the parasites respond to their environment and coordinate intracellular processes are poorly understood. As a part of an effort to understand parasite signaling functions, we report the results of a genome-wide analysis of protein kinases (PKs) of these three trypanosomatids.ResultsBioinformatic searches of the trypanosomatid genomes for eukaryotic PKs (ePKs) and atypical PKs (aPKs) revealed a total of 176 PKs in T. brucei, 190 in T. cruzi and 199 in L. major, most of which are orthologous across the three species. This is approximately 30% of the number in the human host and double that of the malaria parasite, Plasmodium falciparum. The representation of various groups of ePKs differs significantly as compared to humans: trypanosomatids lack receptor-linked tyrosine and tyrosine kinase-like kinases, although they do possess dual-specificity kinases. A relative expansion of the CMGC, STE and NEK groups has occurred. A large number of unique ePKs show no strong affinity to any known group. The trypanosomatids possess few ePKs with predicted transmembrane domains, suggesting that receptor ePKs are rare. Accessory Pfam domains, which are frequently present in human ePKs, are uncommon in trypanosomatid ePKs.ConclusionTrypanosomatids possess a large set of PKs, comprising approximately 2% of each genome, suggesting a key role for phosphorylation in parasite biology. Whilst it was possible to place most of the trypanosomatid ePKs into the seven established groups using bioinformatic analyses, it has not been possible to ascribe function based solely on sequence similarity. Hence the connection of stimuli to protein phosphorylation networks remains enigmatic. The presence of numerous PKs with significant sequence similarity to known drug targets, as well as a large number of unusual kinases that might represent novel targets, strongly argue for functional analysis of these molecules.

Project description:CCCH type zinc finger proteins are RNA binding proteins with regulatory functions at all stages of mRNA metabolism. The best-characterized member, tritetraproline (TTP), binds to AU rich elements in 3' UTRs of unstable mRNAs, mediating their degradation. In kinetoplastids, CCCH type zinc finger proteins have been identified as being involved in the regulation of the life cycle and possibly the cell cycle. To date, no systematic listing of CCCH proteins in kinetoplastids is available.We have identified the complete set of CCCH type zinc finger proteins in the available genomes of the kinetoplastid protozoa Trypanosoma brucei, Trypanosoma cruzi and Leishmania major. One fifths (20%) of all CCCH motifs fall into non-conventional classes and many had not been previously identified. One third of all CCCH proteins have more than one CCCH motif, suggesting multivalent RNA binding. One third have additional recognizable domains. The vast majority are unique to Kinetoplastida or to a subgroup within. Two exceptions are of interest: the putative orthologue of the mRNA nuclear export factor Mex67 and a 3'-5' exoribonuclease restricted to Leishmania species. CCCH motifs are absent from these proteins in other organisms and might be unique, novel features of the Kinetoplastida homologues. Of the others, several have a predicted, and in one case experimentally confirmed, connection to the ubiquitination pathways, for instance a HECT-type E3 ubiquitin ligase. The total number of kinetoplastid CCCH proteins is similar to the number in higher eukaryotes but lower than in yeast. A comparison of the genomic loci between the Trypanosomatidae homologues provides insight into both the evolution of the CCCH proteins as well as the CCCH motifs.This study provides the first systematic listing of the Kinetoplastida CCCH proteins. The number of CCCH proteins with more then one CCCH motif is larger than previously estimated, due to the identification of non-conventional CCCH motifs. Experimental approaches are now necessary to examine the functions of the many unique CCCH proteins as well as the function of the putative Mex67 and the Leishmania 3'-5' exoribonuclease.

Dataset Information

Structures of aspartate aminotransferases from Trypanosoma brucei, Leishmania major and Giardia lamblia.

Publications

Structures of aspartate aminotransferases from Trypanosoma brucei, Leishmania major and Giardia lamblia.

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