Crowding within synaptic junctions influence the degradation of adenoside nucleotides by CD39 and CD73 ectonucleotidases

1 Synapsed cells can communicate using exocytosed nucleotides like adenosine triphosphate (ATP). Ectonucleotidases localized to a synaptic junction degrade such nucleotides into metabolites like adenosine monophosphate (AMP) or adenosine, oftentimes in a sequential manner. CD39 and CD73 are a representative set of coupled ectonucleotidases, where CD39 first converts ATP and adenosine diphosphate (ADP) into AMP, after which the AMP product is dephosphorylated into adenosine by CD73. Hence, CD39/CD73 help shape cellular responses to extracellular ATP. In a previous study [1] we demonstrated that the rates of coupled CD39/CD73 activity within synapse-like junctions are strongly controlled by the enzymes’ co-localization, their surface charge densities, and the electrostatic potential of the surrounding cell membranes. In this study, we demonstrate that crowders within a synaptic junction, which can include globular proteins like cytokines and membrane-bound proteins, impact coupled CD39/CD73 electronucleotidase activity and in turn, the availability of intrasynapse ATP. Specifically, we simulated a spatially-explicit, reaction-diffusion model for the coupled conversion of ATP→AMP and AMP→adenosine in a model synaptic junction with crowders via the finite element method. Our modeling results suggest that the association rate for ATP to CD39 is strongly influenced by the density of intrasynaptic protein crowders, as increasing crowder density suppressed ATP association kinetics. Much of this suppression can be rationalized based on a loss of configurational entropy. The surface charges of crowders can further influence the association rate, with the surprising result that favorable crowder/nucleotide electrostatic interactions can yield CD39 association rates that are faster than crowder-free configurations. However, attractive crowder/nucleotide interactions decrease the rate and efficiency of adenosine production, which in turn increases the availability of ATP and AMP within the synapse relative to crowder-free configurations. These findings highlight how CD39/CD73 ectonucleotidase activity, electrostatics and crowding within synapses influence the availability of nucleotides for intercellular communication.