The topology of the brown adipose tissue mitochondrial uncoupling protein determined with antibodies against its antigenic sites revealed by a library of fusion proteins.
AUTOR(ES)
Miroux, B
RESUMO
The uncoupling protein (UCP) of brown adipose tissue mitochondria is a specialized member of the family of evolutionarily related mitochondrial membrane transporters, which also includes the ADP/ATP translocator and the phosphate carrier. We have generated a library of bacterial clones randomly expressing short subsequences of the UCP fused to the MalE periplasmic protein of Escherichia coli. Anti-UCP sera were used to select clones expressing antigenic sequences of the UCP. Ten different fusion proteins representing eight non-overlapping subsequences of the UCP were obtained. The ability of fusion proteins to select antibodies directed against a short segment of the UCP was used to study the topological organization of the UCP in the inner mitochondrial membrane. Four different fusion proteins were used to determine the orientation of the N-terminal extremities of the first, second, third and fourth predicted alpha-helices of the UCP. This topological study together with previous data on the UCP provides an experimental basis for the predicted structure of the UCP and for other homologous carrier proteins.
ACESSO AO ARTIGO
http://www.pubmedcentral.nih.gov/articlerender.fcgi?artid=413655Documentos Relacionados
- Molecular approach to thermogenesis in brown adipose tissue: cDNA cloning of the mitochondrial uncoupling protein.
- Complete nucleotide and derived amino acid sequence of cDNA encoding the mitochondrial uncoupling protein of rat brown adipose tissue: lack of a mitochondrial targeting presequence.
- Reconstitution of purified brown adipose tissue mitochondria uncoupling protein: demonstration of separate identity of nucleotide binding and proton translocation sites by chemical probes.
- Regulation of mitochondrial protein synthesis by cytoplasmic proteins.
- An upstream enhancer regulating brown-fat-specific expression of the mitochondrial uncoupling protein gene.