Affinity labelling of smooth-muscle myosin light-chain kinase with 5'-[p-(fluorosulphonyl)benzoyl]adenosine - PubMed (original) (raw)
Comparative Study
. 1993 Nov 15;296 ( Pt 1)(Pt 1):53-8.
doi: 10.1042/bj2960053.
Affiliations
- PMID: 8250857
- PMCID: PMC1137654
- DOI: 10.1042/bj2960053
Comparative Study
Affinity labelling of smooth-muscle myosin light-chain kinase with 5'-[p-(fluorosulphonyl)benzoyl]adenosine
H Komatsu et al. Biochem J. 1993.
Abstract
5'-(p-(Fluorosulphonyl)[14C]benzoyl)adenosine (FSBA) was synthesized and used as a probe to study the ATP-binding site of smooth-muscle myosin light-chain kinase (MLCK). FSBA modified both free MLCK and calmodulin/MLCK complex, resulting in inactivation of the kinase activity. Nearly complete protection of the calmodulin/MLCK complex against FSBA modification was obtained by addition of excess ATP whereas MLCK activity alone was lost in a dose-dependent manner even in the presence of excess ATP. These results suggest that FSBA modified ATP-binding sites and ATP-independent sites, and the latter sites are protected by calmodulin binding. The results also suggest that the ATP-binding site is accessible to the nucleotide substrate regardless of calmodulin binding. The FSBA-labelled MLCK was completely proteolysed by alpha-chymotrypsin, and the 14C-labelled peptides were isolated and sequenced. The sequence of the labelled peptide was Ala-Gly-X-Phe, where X is the labelled residue. The sequence was compared with the known MLCK sequence, and the labelled residue was identified as lysine-548, which is located downstream of the GXGXXG motif conserved among ATP-utilizing enzymes.
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References
- Science. 1991 Jul 26;253(5018):407-14 - PubMed
- Science. 1992 Oct 2;258(5079):130-5 - PubMed
- J Biol Chem. 1991 Dec 15;266(35):23936-44 - PubMed
- J Biol Chem. 1976 Aug 10;251(15):4501-13 - PubMed
- Anal Biochem. 1976 May 7;72:248-54 - PubMed
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