Assay of dihydroxyphenylalanine (Dopa) in invertebrate structural proteins

The amino acid, 3,4-L-dihydroxyphenylalanine (dopa), occurs in a variety of insoluble sclerotized structures in molluscs, annelids, coelenterates, and possibly flatworms. Assays for detecting dopa proteins are necessary for developing purification strategies, measuring enzymatic posttranslational tyrosine hydroxylation, and determining the interactions of dopa proteins with other substances. Owing to the importance of dopa in the biosynthesis of catecholamines, the number of available assay methods is staggering and includes colorimetric, fluorometric, polarographic, radiochemical, and electrochemical techniques. This chapter describes several techniques that have proved to be useful and reliable for detecting dopa proteins. These techniques are not necessarily the most sensitive, but they are highly reproducible and for the most part require no more equipment than what is routinely available in protein chemistry laboratories. In addition, methods for isolating dopa and dopa proteins are discussed in this chapter. Dopa can easily be isolated from the hydrolysates of dopa-containing proteins by using a combination of ion-exchange chromatography and gel filtration. These consequently facilitate the quantitation of dopa in completely insoluble proteins.