NAD-preferring malic enzyme: localization, regulation and its potential role in herring (Clupea harengus) sperm cells

Abstract

Herring spermatozoa exhibit a high activity of NAD-preferring malic enzyme (NAD-ME). This enzyme is involved in the generation of NADH or NADPH in the decarboxylation of malate to form pyruvate and requires some divalent cations to express its activity. In order to confirm that NAD-ME isolated from herring sperm cells is localized in mitochondria, we performed immunofluorescent analysis and assayed spectrophotometrically the malic enzyme reaction. Production of polyclonal rabbit antibodies against NAD-ME from herring spermatozoa enabled identification of mitochondrial localization of this enzyme inside herring spermatozoa. The kinetic studies revealed that NAD-ME was competitively inhibited by ATP up to tenfold. Addition of fumarate reversed ATP-dependent inhibition of NAD-ME to 55 % of its maximum activity. The pH-dependent regulation of malic enzyme activity was also examined. Malic enzyme showed maximum activity at pH near 7.0 in all studied conditions. Finally, the role of malic enzyme activity regulation in mitochondria of herring sperm cells was discussed.

Bioenergetics of fish spermatozoa with focus on some herring (Clupea harengus) enzymes

“…Localization of ME in mitochondria is important because these organelles are involved in energy production. In herring spermatozoa, mitochondrial NAD-preferring ME was inhibited by ATP and fumarate reversed this inhibition (Niedźwiecka et al 2017). The physiological concentration of ATP (2–3 mM) in the matrix of respiring mitochondria is sufficient to completely inhibit NAD-preferring ME when ATP is reasonably high, the inhibitory effect of ATP must be overcome.…”

“…2). NAD-preferring ME from herring spermatozoa was localized in the mitochondrion by an antibody raised in rabbit against NAD-preferring ME isolated from herring spermatozoa (Niedźwiecka et al 2017). ATP was found to decrease the rate of NAD reduction of NAD-preferring ME from herring spermatozoa with respect to malate as was shown earlier.…”

“…ATP was found to decrease the rate of NAD reduction of NAD-preferring ME from herring spermatozoa with respect to malate as was shown earlier. Two-millimolar ATP reduces the rate of NADH formation to about 10% compared to control conditions (NAD-preferring ME without inhibitor addition) in the presence of 5 mM malate (Niedźwiecka et al 2017). On the other hand, fumarate was very effective in reversing ATP inhibition of NAD-preferring ME from herring testicular tissue and in the presence of 2 mM fumarate, ATP inhibition was reversed to about 70% of the activity in the absence of ATP (Skorkowski and Storey 1990).…”

“…On the other hand, fumarate was very effective in reversing ATP inhibition of NAD-preferring ME from herring testicular tissue and in the presence of 2 mM fumarate, ATP inhibition was reversed to about 70% of the activity in the absence of ATP (Skorkowski and Storey 1990). Inhibition of NAD-preferring ME could be overcome by fumarate and in consequence could adjust spermatozoa metabolism to changing levels of ATP and fumarate (Niedźwiecka et al 2017). Previously, it has been shown that ATP is an effective inhibitor of both NAD- and NADP-linked reaction by NAD(P)-ME from mammals and fish (Sauer 1973; Lin and Davis 1974; Skorkowski and Storey 1988, 1990; Żołnierowicz et al 1988).…”

Studies on structure-function relationships of acetolactate decarboxylase from Enterobacter cloacae

“…39,40 For NAD(P)+-dependent decarboxylases, such as malic enzymes, a divalent metal ion (Mn 2+ or Mg 2+ ) presented signicant inuences on both catalysis and structural stability. 41 Overall structure of E.c.-ALDC…”