Purification and characterization of a cytosolic thyroid-hormone-binding protein (CTBP) in Xenopus liver.
A variety of cytosolic thyroid-hormone-binding proteins with different characteristics have previously been reported. Here, we first describe the thyroid-hormone-binding characteristics of adult Xenopus liver cytosol, then a novel procedure for purifying cytosolic thyroid-hormone-binding protein (CTBP) from Xenopus liver (xCTBP). The procedure consists of combining preparative isoelectrofocusing, FPLC cation-exchange chromatography, HPLC hydrophobic-interaction chromatography and ultraviolet light cross-linking of 125I-labeled 3,3'5-triiodo-L-thyronine (T3). The isolated xCTBP thus prepared retained all the characteristics of the major thyroid- hormone-(TH)-binding component of the unfractionated cytosol. It is a monomeric protein of approximately 59 kDa with an isoelectric point of 7.0 +/- 0.1, binds T3 with a higher affinity than its analogs with a Kd of approximately 9 nM, and is sensitive to sulfhydryl agents but not to NADPH. In several respects, xCTBP differs from most CTBP-like preparations from other sources described hitherto. Microsequencing of a 23-amino-acid peptide generated from xCTBP by cyanogen bromide digestion revealed 92-100% identity of a 23-amino-acid sequence of several mammalian (amino acids 236-258) and avian (amino acids 245-267) cytosolic aldehyde dehydrogenases (ALDH); xCTBP also exhibited significant similarity of amino acid composition with rat ALDH. This novel finding of sequence identity between a CTBP and ALDH, and the diversity of CTBPs from different sources, suggest that a variety of cytosolic proteins, depending on the species and tissue, can function as thyroid-hormone-binding proteins.
Laboratory of Developmental Biochemistry, National Institute for Medical Research, London, England.
K
K
Yamauchi
J R
JR
Tata
D000595:Amino Acid Sequence; D000818:Animals; D001667:Binding, Competitive; D002352:Carrier Proteins / Q000235:genetics / Q000302:isolation & purification* / Q000378:metabolism; D003600:Cytosol / Q000378:metabolism; D005260:Female; D006801:Humans; D066298:In Vitro Techniques; D007526:Isoelectric Point; D008099:Liver / Q000378:metabolism*; D008297:Male; D008565:Membrane Proteins / Q000235:genetics / Q000302:isolation & purification* / Q000378:metabolism; D008969:Molecular Sequence Data; D008970:Molecular Weight; D017386:Sequence Homology, Amino Acid; D013045:Species Specificity; D013963:Thyroid Hormones; D014284:Triiodothyronine / Q000378:metabolism; D014981:Xenopus; D000097810:Thyroid Hormone-Binding Proteins
D002352:Carrier Proteins; D008565:Membrane Proteins; D013963:Thyroid Hormones; D014284:Triiodothyronine; D000097810:Thyroid Hormone-Binding Proteins