Characterization of a novel cGMP binding protein from rat lung.

Abstract

A specific cGMP binding protein from rat lung has been partially purified and characterized. The binding protein is clearly distinct from the cGMP-dependent protein kinase as determined by several of its characteristics such as chromatographic behavior and spatial requirements for cGMP binding. The protein has a sedimentation coefficient of 7.8 S, a Stokes radius of 5.5 nm, and an estimated M, = 177,000. A cGMP phosphodiesterase activity co-purifies with the binding protein through several steps (ion exchange chromatography, gel filtration, and sucrose density gradient centrifugation) and the possibility exists that these two activities may be functions of the same protein. Several lines of evidence indicate that the cGMP binding site is distinct from the phosphodiesterase catalytic site. First, 1-methyl-3-isobutylxanthine is a competitive inhibitor of cGMP for phosphodiesterase activity, while under similar conditions this compound increases cGMP binding. Other phosphodiesterase inhibitors also increase cGMP binding and these effects are not due to substrate protection. Second, the cGMP analogue 2’-0monobutyryl cGMP is an effective inhibitor of cGMP phosphodiesterase activity, but at the same concentration this compound has no effect on cGMP binding. Third, heat or trypsin treatment of the protein causes loss of cGMP phosphodiesterase activity, while both treatments increase cGMP binding. The characteristics of the cGMP binding protein-phosphodiesterase and its presence in several cell types, ie. sea urchin sperm and rat platelets, which appear to contain very low levels of cGMP-dependent protein kinase, suggest that this protein may have a physiological role in mediating cGMP action.