Molecular cloning and functional expression of a recombinant 72.5 kDa fragment of the 110 kDa regulatory subunit of smooth muscle protein phosphatase 1M
(1995) FEBS Letters — Vol. 377, n° 2, p. 123-127 (1995)
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- Abstract
- We have cloned a partial rat kidney cDNA that encodes a 72.5 kDa N terminal fragment of a third isoform of the M110 subunit of phosphatase 1. This new isoform contains an insert in the 542-597 position not present in the M110 previously cloned (Chen et al. (1994) FEBS Lett. 356, 51-55) from the same species. The encoded cDNA was expressed as a soluble GST-fusion protein in E. coli, and its ability to interact with native PP-1C was measured both in vitro and in permeabilized smooth muscle. In vitro, the fusion protein was capable of selectively binding PP-1C and increasing the substrate specificity of the phosphatase towards myosin 13.2 +/- 3.5-fold (S.E. of the mean, n = 3). In permeabilized smooth muscle pretreated with microcystin, the recombinant protein alone (1.0 microM) did not cause relaxation, but did significantly enhance the ability of PP-1C (0.3 microM) to relax the muscle. These findings show that the N terminal domain of the M110 subunit is the primary site for both PP-1C and myosin binding, and thereby determines myosin specificity. The presence of isoformic variation within this sequence may permit organ/cell specific regulation of phosphorylation sites.
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Haystead, C. M. M., Gailly, P., Somlyo, A. P., Somlyo, A. V., & Haystead, T. A. J. (1995). Molecular cloning and functional expression of a recombinant 72.5 kDa fragment of the 110 kDa regulatory subunit of smooth muscle protein phosphatase 1M. FEBS Letters, 377(2), 123-127. https://doi.org/10.1016/0014-5793(95)01318-0 (Original work published 1995)