Understanding the mechanisms behind host cell invasion by Plasmodium falciparum parasites remains a major hurdle to developing new effective therapeutics that target the asexual cycle, and symptomatic stage of malaria. Whilst an important role for Ca2+ signalling has been demonstrated in regulating invasion and egress of P. falciparum parasites, less understood is the role of cyclic nucleotide signalling in these processes and how these two pathways are related. An important downstream effector of the cAMP-signalling pathway is Protein Kinase A (PKA), a cAMP-dependent protein kinase. PKA has been previously implicated in parasite invasion; however, the role it plays in infection is poorly understood. In the present study, CRISPR-Cas9 and conditional gene deletion (dimerizable cre) technologies have been implemented to generate a P. falciparum parasite line in which expression of PfPKA is under conditional control. Parasites lacking PKA show severely reduced growth after one intraerythrocytic growth cycle, and live imaging experiments pinpoint an invasion defect following initial attachment. Quantitative phosphoproteomic analysis of parasites following PKA depletion has shown a surprising enrichment of phosphorylation events, particularly in genes associated with Ca2+ signalling. This work not only identifies an essential role for PKA in the P. falciparum asexual life cycle, but demonstrates a potential role for PKA as a negative regulator of Ca2+-dependent phosphorylation, thus providing an understanding of how this kinase functions to regulate erythrocyte invasion.