+ Site Statistics
+ Search Articles
+ Subscribe to Site Feeds
EurekaMag Most Shared ContentMost Shared
EurekaMag PDF Full Text ContentPDF Full Text
+ PDF Full Text
Request PDF Full TextRequest PDF Full Text
+ Follow Us
Follow on FacebookFollow on Facebook
Follow on TwitterFollow on Twitter
Follow on LinkedInFollow on LinkedIn

+ Translate

Specific labeling of the bovine heart mitochondrial phosphate carrier with fluorescein 5-isothiocyanate: roles of Lys185 and putative adenine nucleotide recognition site in phosphate transport

Specific labeling of the bovine heart mitochondrial phosphate carrier with fluorescein 5-isothiocyanate: roles of Lys185 and putative adenine nucleotide recognition site in phosphate transport

Journal of Biological Chemistry 276(13): 9792-9799

The amine/SH-modifying fluorescein 5-isothiocyanate (FITC) specifically labeled Lys(185) in the putative membrane-spanning region of the phosphate carrier from both the cytosolic and matrix sides of bovine heart mitochondria at 0 degrees C and pH 7.2, and the labeling inhibited the phosphate transport. Nonmodifying fluorescein derivatives having similar structural features to those of ADP and ATP (Majima, E., Yamaguchi, N., Chuman, H., Shinohara, Y., Ishida, M., Goto, S., and Terada, H. (1998) Biochemistry 37, 424-432) inhibited the specific FITC labeling and phosphate transport, but the nonfluorescein phenylisothiocyanate did not inhibit FITC labeling, suggesting that there is a region recognizing the adenine nucleotides in the phosphate carrier and that this region is closely associated with the transport activity. The phosphate transport inhibitor pyridoxal 5'-phosphate inhibited the specific FITC labeling, possibly due to competitive modification of Lys(185). In addition, FITC inhibited the ADP transport and specific labeling of the ADP/ATP carrier with the fluorescein SH reagent eosin 5-maleimide. Based on these results, we discuss the structural features of the phosphate carrier in relation to its transport activity.

(PDF emailed within 0-6 h: $19.90)

Accession: 011385186

Download citation: RISBibTeXText

PMID: 11133984

DOI: 10.1074/jbc.M007222200

Related references

Phosphate-induced efflux of adenine nucleotides from rat-heart mitochondria: evaluation of the roles of the phosphate/hydroxyl exchanger and the dicarboxylate carrier. Biochimica et Biophysica Acta 893(3): 470-479, 1987

The recognition of 2 specific binding sites of the adenine nucleotide translocase by palmitoyl coenzyme a in bovine heart mitochondria and sub mitochondrial particles. Biochemical & Biophysical Research Communications 89(3): 837-844, 1979

Binding of the fluorescein derivative eosin Y to the mitochondrial ADP/ATP carrier: characterization of the adenine nucleotide binding site. Biochemistry 37(1): 424-432, 1998

Labeling of sarcoplasmic reticulum peptides with 32P-phosphate and fluorescein 5'-isothiocyanate. Biochemical Pharmacology 35(24): 4571-4573, 1986

The mitochondrial anion transport system--the phosphate and the adenine nucleotide carriers. Ceskoslovenska Fysiologie 36(1): 41-59, 1987

Oligo nucleotide binding site of ribosomal rna 6 methyl adenine methylase of escherichia coli k 12 synthetic poly ribo nucleotide methylation and guanosine 5 di phosphate 3 di phosphate modulation. Federation Proceedings 38(3 PART 1): 229, 1979

Labeling of sarcoplasmic reticulum peptides with phosphorus 32 phosphate and fluorescein 5 isothiocyanate. 1986

Photoaffinity labeling of the mitochondrial phosphate carrier by 4-azido-2-nitrophenyl phosphate. Biochimica et Biophysica Acta 890(1): 39-46, 1987

Phosphate stimulation of electrogenic adenine nucleotide translocation mediated by mitochondrial phosphate transporter. Israel Journal of Medical Sciences 13(9): 950, 1977

The roles of phosphate and the phosphate carrier in the mitochondrial permeability transition pore. Mitochondrion 12(1): 120-125, 2012

Labeling of lysine 492 with pyridoxal 5'-phosphate in the sarcoplasmic reticulum Ca-2+-ATPase. Lysine 492 residue is located outside the fluorescein S-isothiocyanate-binding region in or near the ATP binding site. Journal of Biological Chemistry 268(28): 20930-20936, 1993

Affinity labeling of adenine nucleotide related enzymes with reactive adenine nucleotide analogs part 1 affinity labeling of glyceraldehyde 3 phosphate dehydrogenase and myo kinase with a reactive amp analog. Journal of Biochemistry (Tokyo) 81(4): 1147-1154, 1977

Affinity labeling of adenine nucleotide-related enzymes with reactive adenine nucleotide analogs. I. Affinity labeling of glyceraldehyde 3-phosphate dehydrogenase and myokinase with a reactive AMP analog. Journal of Biochemistry 81(4): 1147-1154, 1977

Phosphate sensitive and membrane side specific inhibition of the mitochondrial phosphate transport protein by 4 azido 2 nitrophenyl phosphate. Biophysical Journalpart 2: 183a, 1987