Efflux of amino acids

Although uptake and accumulation of most amino acids from the external medium seems to be irreversible, amino acids are excreted into the medium whenever they are overproduced above a given threshold by yeast cells [6], This can occur under a number of specific conditions, namely in mutants with impaired regulation of amino acid biosynthesis, or in the presence of mutations preventing substrate catabolism, or when growth occurs in the presence of metabolic intermediates. It can even occur when growth is arrested under conditions where amino acid synthesis can continue. 

Except for its narrow specificity, the ATRI gene product shares a number of properties with the higher eukaryotic MDR proteins responsible for multidrug resistance in tumour cells. The MDR gene products are also transmembrane proteins which seem to function as ATP-dependent drug-efflux pumps pumping out a variety of structurally unrelated compounds (see [25,26]). 

A large number of amino acid transporters have been detected by isolating mutations which selectively inactivate one permease without altering enzyme activities involving the corresponding amino acid. Competitive inhibition, kinetics and regulatory behaviour have also been used as criteria to distinguish one transport system from another (see section 4.2). 

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D. L. Jones and P. R. Darrah, Influx and efflux of amino acids from Zea mays L. roots and its implications in the rhizosphere. Plant and Soil 163 (1994). 

The efflux of amino acids from skeletal muscle supports the essential amino acid pool in the blood (see Fig. 42.3). Skeletal muscle oxidizes the BCAA (valine, leucine, isoleucine) to produce energy and glutamine. The amino groups of the BCAA, and of aspartate and glutamate, are transferred out of skeletal muscle in alanine and glutamine. Alanine and glutamine account for approximately 50% of the total a-amino nitrogen released by skeletal muscle (Fig. 42.4). 

When alanine is a major end-product of glucose catabolism, substantial amounts of nitrogenous compounds must be available to contribute the amino group. In a number of species alanine formation is accompanied by proteolysis which would release a pool of amino acids from which the nitrogen could be derived. In G. lamblia and Trichomonas vaginalis arginine catabolism is a likely provider of nitrogen for alanine synthesis. The efflux of alanine from G. lamblia is due to an alanine antiport which is also responsible for alanine uptake and acts to maintain a balance between intracellular and extracellular alanine concentrations (36). 

When the carbon skeleton of alanine is derived from glucose, the efflux of alanine from skeletal muscle and its uptake by liver provide no net transfer of amino acid carbon to the liver for gluconeogenesis. However, some of the alanine carbon is derived from sources other than glucose. Which amino acids can provide carbon for alanine formation (Hint See Fig. 42.9.)... 

In earlier studies, the efflux of exogenously applied, radioactive amino acids was most commonly studied (Katz et al, 1969, Snnivasan et al., 1969). The high sensitivity of the postcolumn denvatization technique with fluorigenic derivatization made it possible to follow the release of a range of endogenous amino acids during various conditions (Norris et al., 1980). Moreover, the precolumn derivatization techniques (Lindroth and Mopper, 1979, Ejnarsson et al, 1983) enable measurement of subpicomol amounts of amino acids. 

The semilogarithmic plot is resolved into components by the method of subtraction, and the half-time (fi/2) of each component is determined graphically. The rate of efflux of labeled solute (k, %/min) is calculated from k = 0.693/fi/2 x 100 (Cutler et al., 1971). This method reveals a fast component for amino acids, as well as other compounds of different molecular weight. It has a ty2 of 2-3 mm in tissue slices for all components studied and probably represents the washout of adherent medium. The slow component is the loss of isotope from the tissue as a whole it is different for different substances and is sensitive to temperature changes The apparent first order kinetics of the slow component does not preclude the possibility that the amino acids are lost from different tissue compartments at different rates. This does, however, suggest that the loss from one major compartment to another may be rate-limiting for clearance from the whole tissue (Cutler et al., 1971). In brain slices, the slow exponential loss of amino acids is linear throughout 40 min of superfusion, and at the end of this time, 60-70% of the labeled amino acids are recovered in the effluent (Cutler et al., 1971)... 

An experimental protocol is described below for monitoring efflux rates of amino acids from cellular preparations. The methodology can be used for a variety of preparations, including homogenates, subcellular fractions, small tissue slices, and cell suspensions The experimental protocol given here is for studying [ C]-GABA efflux from rat brain synaptosomal fractions, as described by Cotman and coworkers (Levy et al., 1973, Redburn et al, 1975, Cotman et al., 1976). 

The amino acid overproduction is also a function of amino acid secreted out of the cell. This can be achieved by overexpressing amino acid carrier proteins which increases amino acid efflux and relieves feedback inhibition. 

Fig. 13.1. The transport of amino acids across the intestinal and tubular cells involves several steps, which are demonstrated schematically for a proximal tubular cell AAy amino acid BLMy basolateral membrane BBMy brush border membrane TBMy tubular basement membrane ly specific transporter at the luminal membrane 2, passive efflux from cytosol into tubular lumen 3, transport system at the basolateral membrane 4, transport from peritubular site into cytosol 5, energy production for electrolyte transport 5, passive efflux via paracellular gaps. (Taken from Foreman, JW and Segal, S (1987) Fanconi syndrome. In Pediatric Nephrology (eds. Holliday MA, Barratt TM and Vernier RC), pp 547-565, Williams and Wilkins, Baltimore, with permission to and modified by Brodehl J)...

Leuchtenbeiger W, Huthmacher K, Drauz K (2005) Biotechnological production of amino acids and derivatives current status and prospects. Appl Microbiol Biotechnol 69 1-8 Liu JQ, Dairi T, Itoh N, Kataoka M, Shimizu S, Yamada H (1998) Gene cloning, biochemical characterization and physiological characterization of a thermostable low-specificity L-threonine aldolase from Escherichia coli. Eur J Biochem 255 220-226 Livshits VA, Zakataeva NP, Aleshin VV, Vitushkina MV (2003) Identification and characterization of the new gene rhtA involved in threonine and homoserine efflux in Escherichia coli. Res Microbiol 154 123-135... 

Another early effect of growth hormone on skeletal muscle which can be detected following either the injection of the hormone (8,9) or its addition to medium bathing isolated muscle (2,10,11), is the stimulation of the net transport of certain amino acids into the intracellular compartment of the tissue. Recent studies (4) using the isolated rat diaphragm and the nonutilizable amino acid, a-aminoisobutyric acid (AIB), have indicated that the stimulatory effect of growth hormone on the transport mechanism is due to the enhancement of amino acid influx rather than to the retardation of amino acid efflux from the cells. The effect is not immediate but becomes evident at about the time (20 minutes of incubation) when a significant stimulation of protein synthesis has occurred (4). 

Sialic acid residues were found to be important in the transport of amino acids and proteins in cancer cells, since neuraminidase treatment of HeLa cells decreased the net accumulation of a-aminoisobutyric acid without altering the rate of efflux of preloaded cells (Brown and Michael, 1969), and the same treatment of L1210 leukemia cells inhibited the outward flow of proteins without influencing lysis or the release of nucleosides or sugars. Some relative specificity in the release of proteins was shown by disk-gel electrophoresis (Click et aL, 1966). 

Hu, M. and R. Borchardt. Transport of a large neutral amino acid in a human intestinal epithelial cell line (Caco-2) uptake and efflux of phenylalanine., Biochim. Biophys. Acta 1992, 3335, 233-244... 

The Major Facilitator Superfamily (MFS) [95-97] is the largest secondary transporter family known in the genomes sequenced to date [98], These polytopic integral membrane proteins enable the transport of a wide range of solutes, including amino acids, sugars, ions, and toxins. Medically relevant members of the family include the bacterial efflux pumps associated with... 

The fundamental process of transferring a solute from the environment, across a cell(s) to the blood involves the common steps of adsorption, influx, intracellular trafficking or distribution, and efflux to the blood. This process applies to physiologically important solutes like Na+ and amino acids, and applies equally to solutes of environmental concern, such as toxic metals. Animals spend a significant portion of their energy budgets on osmotic regulation indeed, at the... 

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