TABLE 1.

Reactions catalyzed by NfnAB from Clostridium kluyveri after FeS cluster reconstitutiona

ReactionApparent Kmh (mM)Apparent Vmax (U/mg)
NADPHb→TTC (NAD+ stimulated)0.001 (NADPH)2 (− NAD+)
0.01 (NAD+)8 (+ NAD+)
NADPHb→Fdox (NAD+ dependent)0.025 (NADPH)0.1 (− NAD+)
1e (NAD+)23 (+ NAD+)
NADPHb→NAD+ (Fdoxc dependent)0.001 (NADPH)<0.1 (− Fd)
2 (NAD+)8 (+ Fdoxc)
4.4 (+ Fdredd)
Fdredd→NADP+ (NADH dependent)0.1 (NADP+)0.2 (− NADH)
0.01 (NADH)28 (+ NADH)
<0.01 (− Fd)
NADH→NADP+ (Fdredd dependent)0.01 (NADH)<0.01 (− Fd)
0.1 (NADP+)28 (+ Fdredd)
<0.01 (+ Fdoxc)
NADPHb→benzyl viologen0.001 (NADPH)15 (− NAD+)
0.015 (benzyl viologen)24 (+ NAD+)f
NADH→benzyl viologen0.6 (− NADP+)
0.1 (+ NADP+)g
NADPH→O25.6 (− NAD+)
20 (+ NAD+)i
NADH→O21 (− NADP+)
0.4 (+ NADP+)
  • a Ferredoxin was present only where indicated.

  • b NADPH regeneration system (glucose-6-phosphate dehydrogenase and glucose-6-phosphate).

  • c Fdox regeneration system (hydrogenase and 100% N2).

  • d Fdred regeneration system (hydrogenase and 100% H2), keeping the ferredoxin about 50% reduced.

  • e An apparent Km for NAD+ of 0.1 mM was determined for cell extracts of C. kluyveri (34).

  • f 2 mM NAD+.

  • g 2 mM NADP+.

  • h Extrapolated from Lineweaver-Burk plots and rounded.

  • i NAD+ regeneration system (lactate dehydrogenase and pyruvate).