Recombinant expression and functional characterization of a G3-family protein from Arabidopsis thaliana

Abstract

Adenylate cyclases (ACs) are a diverse group of enzymes, which catalyze the formation of 3ʹ,5ʹ cyclic adenosine monophosphate (cAMP) from adenosine 5ʹ-triphophate (ATP). Cyclic AMP is a key secondary messenger known to be involved in the mediation of responses to various extracellular stimuli in nearly all living organisms. Recently, a total of 14 candidate AC-encoding proteins in the Arabidopsis thaliana genome were identified and proposed through a bioinformatics approach based on the functionally assigned amino acids residues in the catalytic centre of annotated nucleotide cyclases. Among these identified candidates, is a G-3 family protein (G-3FP), which harbours a novel adenylate cyclase catalytic centre and is encoded by the At2g11890 Arabidopsis gene. Incidentally, to date, there are only 4 annotated and experimentally confirmed ACs in plants, which are the Zea mays pollen protein (Moutinho et al., 2001), the Arabidopsis thaliana pentatricopeptide repeat protein (Ruzvidzo et al., 2013), the Nicotiana benthamiana adenylyl cyclase protein (Ito et al., 2014) and the Hippeastrum hybridum adenylyl cyclase protein (Świeżawska et al., 2014). Therefore, with a view to attempt and identify other additional higher plant AC candidates, this study was set out to clone, partially express and functionally characterize the annotated G-3FP protein. Findings from this approach unequivocally demonstrated that this putative protein candidate is indeed a bona fide functional higher plant AC molecule.