Tryptophan 224 of the rat mitochondrial carnitine/acylcarnitine carrier is crucial for the antiport mechanism

Publication date: Available online 21 July 2019Source: Biochimica et Biophysica Acta (BBA) - BioenergeticsAuthor(s): Nicola Giangregorio, Annamaria Tonazzi, Lara Console, Mariella Pistillo, Vito Scalera, Cesare IndiveriAbstractThe mitochondrial carnitine/acylcarnitine carrier (CACT) catalyzes an antiport of carnitine and acylcarnitines and also a uniport reaction with a rate of about one tenth with respect to the antiport rate. The antiport process results from the coupling of the two uniport reactions in opposite directions. In this mechanism, the transition of the carrier from the outward open conformation to the inward open one (or vice versa) is much faster for the carrier-substrate complex than for the unbound carrier. To investigate the molecular determinants that couple the binding of the substrate with the conformational transitions, site directed mutagenesis has been employed. The antiport or the uniport reaction was followed as [3H]carnitine uptake in or efflux from proteoliposomes reconstituted with the WT or Trp mutants of the rat CACT. Substitution of each the three Trp residues led to different results. Nearly no variations were observed upon substitution of W192 and/or W296 with Ala. While, substantial alteration of the transport function was observed in the mutants W224A, W224Y and W224F. Mutation of W224 led to the loss of the antiport function while the uniport function was unaltered. In these mutants impairment of the substrate affinity on the external side...

Source: Biochimica et Biophysica Acta (BBA) Bioenergetics - July 22, 2019 Category: Biochemistry Source Type: research