(1) N-Ethylmaleimide (a penetrating SH- reagent) inactivated L-[14C]leucine entrance (binding and translocation) into Saccharomyces cerevisiae, the extent of inhibition depending on the time of preincubation with N-ethylmaleimide, N-ethylmaleimide concentration, the amino acid external and internal concentration, and the energization state of the yeast cells. With D-glucose-energized yeast, N-ethylmaleimide inhibited L-[14C]leucine entrance in all the assayed experimental conditions, but with starved yeast and low (0.1 mM) amino acid concentration, it did not inhibit L-[14C]leucine binding, except when the cells were preincubated with L-leucine. With the rho- respiratory-deficient mutant (energized cells), N-ethylmaleimide inhibited L-[14C]leucine entrance as with the energized wild-type, though to a lesser extent. (2) Analysis of the N-ethylmaleimide effect as a function of L-[14C]leucine concentration showed a significant decrease of Jmax values of the high- (S1) and low- (S2) affinity amino acid transport systems, but KT values were not significantly modified. (3) When assayed in the presence of D-glucose, N-ethylmaleimide inhibition of D-glucose uptake and respiration contributed significantly to inactivation of L-[14C]leucine entrance. Pretreatment of yeast cells with 2,4-dinitrophenol enhanced the effect of L-[14C]leucine binding and translocation. (4) Bromoacetylsulfanilic acid and bromoacetylaminoisophthalic acid, two non-penetrating SH- reagents, did not inactivate L-[14C]leucine entrance, while p-chloromercuribenzoate, a slowly penetrating SH-reagent, inactivated it to a limited extent. When compared with the effect of N-ethylmaleimide, these negative results indicate that thiol groups of the L-[14C]leucine carrier were not exposed on the outer surface of the yeast cell permeability barrier.
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| Evidence ID | Analyze ID | Gene/Complex | Systematic Name/Complex Accession | Qualifier | Gene Ontology Term ID | Gene Ontology Term | Aspect | Annotation Extension | Evidence | Method | Source | Assigned On | Reference |
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| Evidence ID | Analyze ID | Gene | Gene Systematic Name | Phenotype | Experiment Type | Experiment Type Category | Mutant Information | Strain Background | Chemical | Details | Reference |
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| Evidence ID | Analyze ID | Gene | Gene Systematic Name | Disease Ontology Term | Disease Ontology Term ID | Qualifier | Evidence | Method | Source | Assigned On | Reference |
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| Evidence ID | Analyze ID | Regulator | Regulator Systematic Name | Target | Target Systematic Name | Direction | Regulation of | Happens During | Regulator Type | Direction | Regulation Of | Happens During | Method | Evidence | Strain Background | Reference |
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| Site | Modification | Modifier | Source | Reference |
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| Evidence ID | Analyze ID | Interactor | Interactor Systematic Name | Interactor | Interactor Systematic Name | Allele | Assay | Annotation | Action | Phenotype | SGA score | P-value | Source | Reference | Note |
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| Evidence ID | Analyze ID | Interactor | Interactor Systematic Name | Interactor | Interactor Systematic Name | Assay | Annotation | Action | Modification | Source | Reference | Note |
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| Complement ID | Locus ID | Gene | Species | Gene ID | Strain background | Direction | Details | Source | Reference |
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| Evidence ID | Analyze ID | Dataset | Description | Keywords | Number of Conditions | Reference |
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