Riboflavin binding protein, copper and phosvitin: Thermodynamic and electrochemical investigations of protein-metal and protein-protein interactions

Riboflavin Binding Protein (RBP) is a critical transport protein in oviparous species, responsible for the delivery of the essential vitamin riboflavin to the oocyte. We hypothesize the involvement of copper, either in this transport system or independent of it, based on the observation that RBP binds copper in vitro. The equilibrium binding constants of proteins to metal ions can often be used to infer the role of the protein or the metal in biological interactions. Determination of the binding constant of RBP for copper is complicated by the simultaneous reduction of the copper upon binding. Ongoing efforts in our lab to deconvolute the binding event using a combination of electrochemical and physical measurements will be presented. In particular, we have applied isothermal calorimetry (ITC) and cyclic voltammetry (CV) to the problem.

While RBP is known to be a transporter for riboflavin, RBP itself only gains access to the oocyte via a receptor for phosvitin. Phosvitin is part of the primary lipid transport system to eggs, composed of vitellogenin and its constituent proteins lipovitellin and phosvitin. These three proteins are also known to bind metal ions in the egg. Phosvitin and RBP are known to interact at the receptor, possibly facilitating copper exchange. Therefore, we further explore the interactions between RBP, phosvitin, and copper by the aforementioned techniques.