HOCGO AND DMACGO - 2 COUMARIN DERIVED ALPHA-DICARBONYLS SUITABLE AS PH AND POLARITY SENSITIVE FLUORESCENT REPORTERS FOR PROTEINS THAT CAN BE TARGETED AT REACTIVE ARGININES

Two coumarin derived alpha-dicarbonyls, HOCGO and DMACGO, are presente d as pH, polarity and quencher sensitive fluorescent reporters for pro teins that can be targeted at reactive arginines. Both inactivate a nu mber of enzymes that feature functionally critical and chemically susc eptible arginyls. Both are chemoselective in responding towards Arg si de chain but not towards Cys or Lys side chains under suitably dilute conditions. With pK(app) approximate to 6.7, HOCGO can serve as a pH s ensor, while with pK(app) much less than 4.0, DMACGO is better suited as a polarity sensor. A contrasting set of changes are manifest in the CGOs upon protein interaction that are either attributable to Arg mod ification or to the noncovalent probe associations with hydrophobic pr otein domains. DMACGO probes a single hydrophobic site on ovalbumin wh ile HOCGO is largely unresponsive to this protein. Three to five argin yls are modified in HSA and BSA by HOCGO as well as DMACGO, while the latter also probes two hydrophobic sites on both these proteins. HOCGO modifies a single arginine in LDH active site, while its adducts with H-4 and M(4)LDH isozymes titrate to the apparent pK(a) of 7.8. Other proteins labeled with HOCGO or DMACGO reveal a number of variations th at can furnish information about the microenvironment in the sites pro bed. The CGOs are thus potentially useful reporters of protein domains that feature reactive arginines. Suitable experimental condition are defined based on mechanistic considerations that may be used in applyi ng the CGOs as Arg modifiers and as fluorescent probes.