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Amino Acid Permease
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Chris A Kaiser – One of the best experts on this subject based on the ideXlab platform.
transport activity dependent intracellular sorting of the yeast general Amino Acid PermeaseMolecular Biology of the Cell, 2011Co-Authors: Natalie E Cain, Chris A KaiserAbstract:
Intracellular trafficking of the general Amino Acid Permease, Gap1p, of Saccharomyces cerevisiae is regulated by Amino Acid abundance. When Amino Acids are scarce Gap1p is sorted to the plasma membrane, whereas when Amino Acids are abundant Gap1p is sorted from the trans-Golgi through the multivesicular endosome (MVE) and to the vacuole. Here we test the hypothesis that Gap1p itself is the sensor of Amino Acid abundance by examining the trafficking of Gap1p mutants with altered substrate specificity and transport activity. We show that trafficking of mutant Gap1pA297V, which does not transport basic Amino Acids, is also not regulated by these Amino Acids. Furthermore, we have identified a catalytically inactive mutant that does not respond to complex Amino Acid mixtures and constitutively sorts Gap1p to the plasma membrane. Previously we showed that Amino Acids govern the propensity of Gap1p to recycle from the MVE to the plasma membrane. Here we propose that in the presence of substrate the steady-state conformation of Gap1p shifts to a state that is unable to be recycled from the MVE. These results indicate a parsimonious regulatory mechanism by which Gap1p senses its transport substrates to set an appropriate level of transporter activity at the cell surface.
activity dependent reversible inactivation of the general Amino Acid PermeaseMolecular Biology of the Cell, 2006Co-Authors: April L Risinger, Natalie E Cain, Esther J Chen, Chris A KaiserAbstract:
The general Amino Acid Permease, Gap1p, of Saccharomyces cerevisiae transports all naturally occurring Amino Acids into yeast cells for use as a nitrogen source. Previous studies have shown that a …
Activity-dependent reversible inactivation of the general Amino Acid Permease.Molecular Biology of the Cell, 2006Co-Authors: April L Risinger, Natalie E Cain, Esther J Chen, Chris A KaiserAbstract:
: The general Amino Acid Permease, Gap1p, of Saccharomyces cerevisiae transports all naturally occurring Amino Acids into yeast cells for use as a nitrogen source. Previous studies have shown that a nonubiquitinateable form of the Permease, Gap1p(K9R,K16R), is constitutively localized to the plasma membrane. Here, we report that Amino Acid transport activity of Gap1p(K9R,K16R) can be rapidly and reversibly inactivated at the plasma membrane by the presence of Amino Acid mixtures. Surprisingly, we also find that addition of most single Amino Acids is lethal to Gap1p(K9R,K16R)-expressing cells, whereas mixtures of Amino Acids are less toxic. This toxicity appears to be the consequence of uptake of unusually large quantities of a single Amino Acid. Exploiting this toxicity, we isolated gap1 alleles deficient in transport of a subset of Amino Acids. Using these mutations, we show that Gap1p inactivation at the plasma membrane does not depend on the presence of either extracellular or intracellular Amino Acids, but does require active Amino Acid transport by Gap1p. Together, our findings uncover a new mechanism for inhibition of Permease activity in response to elevated Amino Acid levels and provide a physiological explanation for the stringent regulation of Gap1p activity in response to Amino Acids.
Philip S Poole – One of the best experts on this subject based on the ideXlab platform.
rhizobium leguminosarum has a second general Amino Acid Permease with unusually broad substrate specificity and high similarity to branched chain Amino Acid transporters bra liv of the abc familyJournal of Bacteriology, 2002Co-Authors: Arthur H F Hosie, David Allaway, C S Galloway, H A Dunsby, Philip S PooleAbstract:
Amino Acid uptake by Rhizobium leguminosarum is dominated by two ABC transporters, the general Amino Acid Permease (Aap) and the branched-chain Amino Acid Permease (BraRl). Characterization of the solute specificity of BraRl shows it to be the second general Amino Acid Permease of R. leguminosarum. Although BraRl has high sequence identity to members of the family of hydrophobic Amino Acid transporters (HAAT), it transports a broad range of solutes, including Acidic and basic polar Amino Acids (l-glutamate, l-arginine, and l-histidine), in addition to neutral Amino Acids (l-alanine and l-leucine). While Amino and carboxyl groups are required for transport, solutes do not have to be α-Amino Acids. Consistent with this, BraRl is the first ABC transporter to be shown to transport γ-Aminobutyric Acid (GABA). All previously identified bacterial GABA transporters are secondary carriers of the Amino Acid-polyamine-organocation (APC) superfamily. Also, transport by BraRl does not appear to be stereospecific as d Amino Acids cause significant inhibition of uptake of l-glutamate and l-leucine. Unlike all other solutes tested, l-alanine uptake is not dependent on solute binding protein BraCRl. Therefore, a second, unidentified solute binding protein may interact with the BraDEFGRl membrane complex during l-alanine uptake. Overall, the data indicate that BraRl is a general Amino Acid Permease of the HAAT family. Furthermore, BraRl has the broadest solute specificity of any characterized bacterial Amino Acid transporter.
the general Amino Acid Permease of rhizobium leguminosarum strain 3841 is negatively regulated by the ntr systemFems Microbiology Letters, 1997Co-Authors: D L Walshaw, Colm J Reid, Philip S PooleAbstract:
Cosmid-borne and chromosomal lacZ fusions to aapJ, aapQ and aapM were used to examine the nitrogen regulation of the general Amino Acid Permease (Aap) of Rhizobium leguminosarum strain 3841. Transcription of the first gene of the operon (aapJ), which encodes the periplasmic binding protein, was 2–4-fold higher than aapQ and aapM, which encode the integral membrane proteins, under various growth conditions. This may be due to the presence of a putative stem loop in the intergenic region between aapJ and aapQ. All aap fusions were derepressed 3–5-fold after growth on glutamate as a nitrogen source, which effectively causes nitrogen limitation. An ntrC mutant was derepressed for transcription of the aap operon and had high rates of Amino Acid transport when grown on ammonia as the nitrogen source. Thus NtrC negatively regulates the aap operon, contrary to its usual role in positive gene activation. These results confirm that the aap operon is subject to complex regulation involving both transcriptional and post-transcriptional factors.
regulation of the tca cycle and the general Amino Acid Permease by overflow metabolism in rhizobium leguminosarumMicrobiology, 1997Co-Authors: D L Walshaw, Adam Wilkinson, Mathius Mundy, Mary Smith, Philip S PooleAbstract:
Summary: Mutants of Rhizobium leguminosarum were selected that were altered in the uptake activity of the general Amino Acid Permease (Aap). The main class of mutant maps to sucA and sucD, which are part of a gene cluster mdh-sucCDAB, which codes for malate dehydrogenase (mdh), succinyl-CoA synthetase (sucCD) and components of the 2-oxoglutarate dehydrogenase complex (sucAB). Mutation of either sucC or sucD prevents expression of 2-oxoglutarate dehydrogenase (sucAB). Conversely, mutation of sucA or sucB results in much higher levels of succinyl-CoA synthetase and malate dehydrogenase activity. These results suggest that the genes mdh-sucCDAB may constitute an operon. suc mutants, unlike the wild-type, excrete large quantities of glutamate and 2-oxoglutarate. Concomitant with mutation of sucA or sucD, the intracellular concentration of glutamate but not 2-oxoglutarate was highly elevated, suggesting that 2-oxoglutarate normally feeds into the glutamate pool. Elevation of the intracellular glutamate pool appeared to be coupled to glutamate excretion as part of an overflow pathway for regulation of the TCA cycle. Amino Acid uptake via the Aap of R. leguminosarum was strongly inhibited in the suc mutants, even though the transcription level of the aap operon was the same as the wild-type. This is consistent with previous observations that the Aap, which influences glutamate excretion in R. leguminosarum, has uptake inhibited when excretion occurs. Another class of mutant impaired in uptake by the Aap is mutated in polyhydroxybutyrate synthase (phaC). Mutants of succinyl-CoA synthetase (sucD) or 2-oxoglutarate dehydrogenase (sucA) form ineffective nodules. However, mutants of aap, which are unable to grow on glutamate as a carbon source in laboratory culture, show wild-type levels of nitrogen fixation. This indicates that glutamate is not an important carbon and energy source in the bacteroid. Instead glutamate synthesis, like polyhydroxybutyrate synthesis, appears to be a sink for carbon and recluctant, formed when the 2-oxoglutarate dehydrogenase complex is blocked. This is in accord with previous observations that bacteroids synthesize high concentrations of glutamate. Overall the data show that the TCA cycle in R. leguminosarum is regulated by Amino Acid excretion and polyhydroxybutyrate biosynthesis which act as overflow pathways for excess carbon and reductant.
Tomas Girbes – One of the best experts on this subject based on the ideXlab platform.
Effect of l-azetidine 2-carboxilic Acid on the activity of the general Amino–Acid Permease from Saccharomyces cerevisiae var. ellipsoideus
Archives of Microbiology, 1991Co-Authors: Rosario Iglesias, J M Ferreras, F J Arias, R Munoz, Ma Angeles Rojo, Tomas GirbesAbstract:
Addition of the l -proline analogue l -azetidine 2-carboxylic Acid to growing cultures of Saccharomyces cerevisiae var. ellipsoideus promoted fast deactivation of the general AminoAcid Permease, measured as l -valine uptake, without an immediate decrease in the growth rate. Cells preincubated with the analogue for 3 h were unable to restore either growth ability or general AminoAcid Permease activity in analogue-free medium. Eadie-Hofstee plots of l -valine uptake in the presence of the analogue are consistent with a strong reduction in the number of active molecules of the general Amino–Acid Permease located in the plasma membrane. Inhibitory effects on protein synthesis were seen after preincubations of the yeast with the analogue for 3 h although a 30 min preincubation had no effect.
changes in the activity of the general Amino Acid Permease from saccharomyces cerevisiae var ellipsoideus during fermentationBiotechnology and Bioengineering, 1990Co-Authors: R Lglesias, J M Ferreras, F J Arias, R Munoz, Tomas GirbesAbstract:
: The evolution of the activity of the general Amino Acid Permease and ethanol and glucose concentrations in the medium were studied in a mild fermentation process carried out by a wine strain of Saccharomyces cerevisiae var. ellipsoideus isolated from grape musts in spontaneous fermentation. The cells displayed a reduction in the activity of the general Amino Acid Permease parallel to the increase of ethanol in the medium. This ethanol increase was not enough to promote a substantial inhibition on the total polypeptide synthesis measured as polyuridylic-Acid-directed polyphenylalanine synthesis.
differential d glucose requirements of the general Amino Acid Permease and protein synthesis in saccharomyces cerevisiae var ellipsoideusCellular and Molecular Biology, 1990Co-Authors: Rosario Iglesias, J M Ferreras, F J Arias, R Munoz, M A Rojo, Tomas GirbesAbstract:
: The dependence of the general AminoAcid Permease and protein synthesis on the availability of D-glucose as energy source was studied. Stimulation by the sugar was immediate once added to the cell suspensions and seems to be mediated by energy derived directly from glycolysis. The general AminoAcid Permease was saturated linearly with D-glucose whereas protein synthesis was saturated sigmoidealy requiring much higher concentration of the sugar than the general AminoAcid Permease.