Active Transport of Glutamate in Leishmania (Leishmania) amazonensis

Abstract

Leishmania spp. are the causative agents of leishmaniasis, a complex of diseases with a broad spectrum of clinical manifestations. Leishmania (Leishmania) amazonensis is a main etiological agent of diffuse cutaneous leishmaniasis. Leishmania spp., as other trypanosomatids, possess a metabolism based significantly on the consumption of amino acids. However, the transport of amino acids in these organisms remains poorly understood with few exceptions. Glutamate transport is an important biological process in many organisms. In the present work, the transport of glutamate is characterized. This process is performed by a single kinetic system (K(m)=0.59+/-0.04 mM, V(max)=0.123+/-0.003 nmol/min per 20 x 10(6) cells) showing an energy of activation of 52.38+/-4.7 kJ/mol and was shown to be partially inhibited by analogues, such as glutamine, aspartate, alpha-ketoglutarate and oxaloacetate, methionine, and alanine. The transport activity was sensitive to the extracellular concentration of H(+) but not to Na(+) or K(+). However, unlike other amino acid transporters presently characterized, the treatment with specific ionophores confirmed the participation of a K(+), and not H(+) membrane gradient in the transport process.

Multi-analytical platform metabolomic approach to study miltefosine mechanism of action and resistance in Leishmania

“…Pro, the other precursor for Glu also decreased which could also explain the increased metabolism of Glu. In fact it has been reported that only 30 % of the overall Glu uptake is incorporated in proteins [77], so here it could be involved with α-ketoglutarate to replenish the mitochondrial TCA directly by glutamate dehydrogenase, or by transamination with pyruvate to produce Ala.…”

“…Amino acids, especially Asp in Leishmania, play an important role in metabolism; transaminases convert Glu and Asp (asparagines) to Ala with α-ketoglutarate, oxaloacetate and pyruvate, all of which are substrates for TCA once inside the mitochondria facilitated by a specific mitochondrial transporter or, for oxoglutarate, after their conversion into malate [74]. Glu and Gln underwent a moderate decreases in ST. A specific transporter for glutamate has been described in Leishmania dependent on K + gradients across the membrane [77]; hence if this gradient is collapsed by membrane leakage and/or shortage of bioenergetic fuel for ionic pumps [24], the Glu level will decrease. Pro, the other precursor for Glu also decreased which could also explain the increased metabolism of Glu.…”

Isotopomer Profiling of Leishmania mexicana Promastigotes Reveals Important Roles for Succinate Fermentation and Aspartate Uptake in Tricarboxylic Acid Cycle (TCA) Anaplerosis, Glutamate Synthesis, and Growth

“…Glutamate is the most abundant free amino acid in promastigote stages (data not shown) but is internalized at a low rate under standard culture conditions. Although the Leishmania glutamate transporter has not been identified, uptake experiments suggest that the promastigote glutamate transporter(s) have a high K m value (48). Conversely, a high rate of de novo glutamate synthesis were suggested by the rapid labeling of this amino acid with [U- 13 C]glucose, [U- 13 C]aspartate, [U-13 C]alanine, and H 13 CO 3 Ϫ .…”

“…Given the importance of glutamate for Leishmania growth and pathogenesis, it is curious why this parasite has such a limited capacity to take up this amino acid. Although the Leishmania glutamate transport has not been studied in detail (48), leakage of intracellular pools was clearly detected during rapid growth (Fig. 2).…”

Capillary electrophoresis reveals polyamine metabolism modulation in Leishmania (Leishmania) amazonensis wild‐type and arginase‐knockout mutants under arginine starvation

“…Similarly to other members of Trypanosomatidae family, Leishmania uses amino acids for different metabolic functions, including cellular volume control and as energy sources . l ‐Arginine is an essential amino acid for these organisms, and there are specific membrane transporters for its uptake .…”