Isoforms of Na,K-ATPase in Artemia saline: I. Detection by FITC binding and time course.

Partially purified Na,K-ATPase from whole nauplii at various stages of development, analyzed by SDS-PAGE, reveals a polydisperse beta and two alpha subunits (denoted alpha 1 and alpha 2). In the absence of Ca2+, ATP-inhibitable fluorescein isothiocyanate (FITC) labeling is restricted to the alpha subunit of this enzyme, even in crude naupliar homogenates. The intensity of the alpha-specific fluorescent signal (i.e., the sum of the yield from both alpha isoforms) is proportional to Na,K-ATPase activity during development. FITC-labeled subunits were detected at 8 hr of development prior to the detection of measurable Na,K-ATPase activity. The alpha 2/alpha 1 ratio changed from an initial value of 1.25 to a peak of 1.75 at 32 hr of development, then reverted to a ratio of 1.25 by 42 hr, and remained constant thereafter. Pulse chase studies with 35S-methionine indicated that the developmental increase in enzyme activity is coincident with amino acid incorporation into the alpha subunits, implying that enzyme synthesis is active during enzyme accumulation.