Molecular and functional characterization of two malic enzymes from Leishmania parasites

[Display omitted] •The two isoforms of leishmanial malic enzyme exhibit an identity of about 55%.•Only one of the two isozymes is allosterically activated by l-aspartate.•Both malic enzyme isoforms specifically catalyze NADPH production.•The two isozymes appear to exhibit a species-specific expressi...

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Published in:	Molecular and biochemical parasitology Vol. 219; pp. 67 - 76
Main Authors:	Giordana, Lucila, Sosa, Máximo Hernán, Leroux, Alejandro E., Mendoza, Elkin F. Rodas, Petray, Patricia, Nowicki, Cristina
Format:	Journal Article
Language:	English
Published:	Netherlands Elsevier B.V 01-01-2018
Subjects:	Aspartic Acid - metabolism Binding Sites Cloning, Molecular Coenzymes - metabolism Computational Biology Gene Expression Leishmania Leishmania major - enzymology Leishmania major - genetics Leishmania mexicana - enzymology Leishmania mexicana - genetics Malate Dehydrogenase (NADP+) - chemistry Malate Dehydrogenase (NADP+) - genetics Malate Dehydrogenase (NADP+) - metabolism Malic enzymes Models, Molecular NADP - metabolism NADPH production Protein Multimerization Redox metabolism Redox metabolism NADPH production Leishmania Malic enzymes
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Summary:	[Display omitted] •The two isoforms of leishmanial malic enzyme exhibit an identity of about 55%.•Only one of the two isozymes is allosterically activated by l-aspartate.•Both malic enzyme isoforms specifically catalyze NADPH production.•The two isozymes appear to exhibit a species-specific expression pattern. Leishmania parasites cause a broad spectrum of clinical manifestations in humans and the available clinical treatments are far from satisfactory. Since these pathogens require large amounts of NADPH to maintain intracellular redox homeostasis, oxidoreductases that catalyze the production of NADPH are considered as potential drug targets against these diseases. In the sequenced genomes of most Leishmania spp. two putative malic enzymes (MEs) with an identity of about 55% have been identified. In this work, the ME from L. major (LmjF24.0770, Lmj_ME-70) and its less similar homolog from L. mexicana (LmxM.24.0761, Lmex_ME-61) were cloned and functionally characterized. Both MEs specifically catalyzed NADPH production, but only Lmex_ME-61 was activated by l-aspartate. Unlike the allosterically activated human ME, Lmex_ME-61 exhibited typical hyperbolic curves without any sign of cooperativity in the absence of l-aspartate. Moreover, Lmex_ME-61 and Lmj_ME-70 differ from higher eukaryotic homologs in that they display dimeric instead of tetrameric molecular organization. Homology modeling analysis showed that Lmex_ME-61 and Lmj_ME-70 notably differ in their surface charge distribution; this feature encompasses the coenzyme binding pockets as well. However, in both isozymes, the residues directly involved in the coenzyme binding exhibited a good degree of conservation. Besides, only Lmex_ME-61 and its closest homologs were immunodetected in cell-free extracts from L. mexicana, L. amazonensis and L. braziliensis promastigotes. Our findings provide a first glimpse into the biochemical properties of leishmanial MEs and suggest that MEs could be potentially related to the metabolic differences among the species of Leishmania parasites.
Bibliography:	ObjectType-Article-1 SourceType-Scholarly Journals-1 ObjectType-Feature-2 content type line 23
ISSN:	0166-6851 1872-9428
DOI:	10.1016/j.molbiopara.2017.11.001