Binding of Acrylonitrile to Parvalbumin

A previous study has shown that acrylonitrile (ACN) has a longhalf-life in rainbow trout muscle and that [¹⁴C]ACN appearsto be bound to a 10,000-Da protein in muscle. The labeled proteinwas purified from muscle of trout exposed to [¹⁴C]ACN, separatedon 20% SDS–PAGE, and digested for amino acid analysisand sequence analysis. These studies indicated that the labeledprotein was the Ca2⁺-binding protein parvalbumin. Parvalbuminis an important calcium-binding protein thought to be involvedin the regulation of calcium levels in various parts of thebody ranging from neurons to fast-twitch muscle contractions.To study the reaction between parvalbumin and [¹⁴C]ACN, frogparvalbumin was incubated with [¹⁴C]ACN in vitro under variousconditions. These studies indicated that the maximum labelingoccurred at 1 nmol/nmol parvalbumin and at pH 7. Amino acidanalysis of the labeled protein indicated that the labeled aminoacid was probably histidine, and endoproteinase Glu-C (V-8)digestion studies revealed that the ¹⁴C was in the 1–81amino acid segment of the protein, an area that contains twohistidines.