Purchase this article with an account.
I. V. Peshenko, A. M. Dizhoor; N-Myristoylation in GCAP1 Affects Its Ca(2+) Sensitivity and Interaction With RetGC1. Invest. Ophthalmol. Vis. Sci. 2010;51(13):1091.
Download citation file:
© ARVO (1962-2015); The Authors (2016-present)
Guanylyl cyclase activating proteins (GCAPs) are myristoylated Ca(2+)/Mg(2+)-binding proteins that impart Ca(2+) sensitivity to retinal guanylyl cyclase (RetGC). The purpose of this study was to elucidate the functional role of the N-myrisloyl group in the interactions between GCAP1 and RetGC1.
Myristoylated and non-myristoylated GCAP1, either untagged or GFP-tagged, was produced in E. coli and HEK293 cells and its metal binding properties and the ability to bind and activate recombinant RetGC1 in vitro or in co-transfected HEK293 cells were assessed using methods previously described in [1, 2].
When myristoylation was blocked by expressing the wild type GCAP1 in the absence of N-myristoyl transferase or by introducing a G2A mutation, it demonstrated Ca(2+)- and Mg(2+)-dependent changes in Trp fluorescence similar to myristoylated WT GCAP1. However, while the spectral changes reflecting transition from an apo-form to a RetGC1 activator form were unaffected in the non-myristoylated GCAP1, the Ca(2+) sensitivity of the transition from the RetGC activator into a RetGC inhibitor form was shifted toward higher Ca(2+) concentration range. Consequently, there was also a decrease in Ca(2+)-dependent inhibition of RetGC1 by the non-myristoylated GCAP1. The lack of myrisloylation also had two additional effects on RetGC1 regulation: it decreased the apparent affinities of both Ca(2+)- and Mg(2+)-bound GCAP1 for RetGC1 and reduced the maximal stimulation of RetGC1 by Mg(2+)-bound GCAP1 in vitro. However, GFP-tagged G2A GCAP1 was still able to associate with RetGC1 in HEK293 cells, although less efficiently than myristoylated GFP-tagged WT GCAP1.
The presence of N-myristoyl group in GCAP1 contributes to all three key elements of the cyclase regulation: binding to RetGC, its maximal activation, and the Ca(2+) sensitivity of its inhibition.
 Peshenko & Dizhoor (2006) J. Biol. Chem. 281, 23830-41;  Peshenko et al. (2008) J. Biol. Chem. 283, 21747-57.
This PDF is available to Subscribers Only