Question

Local cellular pH can decrease from one-half to one pH unit depending on a variety of circumstances including ischemia, hypoxia, and inflammation due to lactic acidosis. In addition, metabolic acidosis is often one of the symptoms in complications following surgery. The investigators have proposed that HPRG acts to relieve the acidosis in these circumstances. Propose a model that explains the mechanism of pH regulation by HPRG.

Answer 1

The middle domain of plasma histidine-proline-rich glycoprotein (HPRG) contains unusual tandem pentapeptide repeats and binds heparin and transition metals. Unlike other proteins that interact with heparin via lysine or arginine residues, HPRG relies exclusively on histidine residues for this interaction. To assess the consequences of this unusual requirement, we have studied the interaction between human plasma HPRG and immobilized glycosaminoglycans (GAGs) using resonant mirror biosensor techniques.

HPRG binding to immobilized heparin was strikingly pH-sensitive, producing a titration curve with a midpoint at pH 6.8. There was little binding of HPRG to heparin at physiological pH in the absence of metals, but the interaction was promoted by nanomolar concentrations of free zinc and copper, and its pH dependence was shifted toward alkaline pH by zinc. The affinity of HPRG for various GAGs measured in a competition assay decreased in the following order: heparin > dermatan sulfate > heparan sulfate > chondroitin sulfate A. Binding of HPRG to immobilized dermatan sulfate had a midpoint at pH 6.5, was less influenced by zinc, and exhibited cooperativity. Importantly, plasminogen interacted specifically with GAG-bound HPRG. We tells that HPRG is a physiological pH sensor, interacting with negatively charged GAGs on cell surfaces only when it acquires a net positive charge by protonation and/or metal binding. This provides a mechanism to regulate the function of HPRG (the local pH) and rationalizes the role of its unique, conserved histidine-proline-rich domain. Thus, under conditions of local acidosis (e.g. ischemia or hypoxia), HPRG can co-immobilize plasminogen at the cell surface as well as compete for heparin with other proteins such as antithrombin.