Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: EC:1.5.1.3 (dihydrofolate reductase)
 5,819 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The cytosolic domain of the human mitochondrial protein import receptor, hTom20, has been expressed as a fusion protein with glutathione S-transferase (GST) in bacteria and the purified protein immobilized on Sepharose beads. To discriminate between specific binding of precursor proteins with the receptor and non-specific binding, precursors were recovered as a complex with GST-hTom20 following competitive elution from the beads with reduced glutathione. Here, we describe the specificity of this assay and demonstrate that the cytosolic domain of hTom20 interacts directly with the transcription-translation product of precursor proteins that bear a diverse array of targeting signals. Such proteins include a matrix protein (pODHFR), a polytopic integral protein of the inner membrane (uncoupling protein), a beta-barrel protein of the outer membrane (VDAC/porin) as well as bitopic integral proteins which are inserted into the outer membrane by either an NH2-terminal or COOH-terminal signal anchor sequence (yTom70(1-29)DHFR and Bcl-2, respectively).
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PMID:Human mitochondrial import receptor, Tom20p. Use of glutathione to reveal specific interactions between Tom20-glutathione S-transferase and mitochondrial precursor proteins. 911 86

The uncoupling protein (UCP1) is a carrier protein of the inner mitochondrial membrane spanning the bilayer six times. It does not contain a typical amino-terminal targeting signal and the mechanism of targeting and insertion is unknown. Here we focus on the biogenesis of UCP1 by analysing the import signals contained within the three repeated units of the protein. The amino-terminal third of the protein can mediate insertion into the outer membrane and therefore acts as artificial targeting signal when fused to DHFR. However, in the context of full-length UCP, the targeting information contained within the first repeated unit is not sufficient to trigger insertion into the outer membrane. Deletion of either the first or third repeated unit from UCP1 did not reduce import into the inner membrane and bound to the outer membrane receptor protein hTom20 with the characteristics of full-length UCP1. Deletion of the second repeat of UCP1 completely abolished all import into the mitochondria. Consistent with this, the central repeat alone was efficiently imported to the inner membrane and bound hTom20 with the characteristics of UCP1. We conclude that the site for binding hTom20 is within the central repeat and that this domain contains the complete targeting signal for directing UCP1 to the inner membrane.
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PMID:The central matrix loop drives import of uncoupling protein 1 into mitochondria. 1082 98