Nitrogenase acetylene reduction

Acetylene-reduction assay Estimates nitrogenase activity by measuring the rate of acetylene reduced to ethylene. 

Substrate reduction by the iron nitrogenase is very similar to that observed with vanadium nitrogenases. Acetylene is a relatively poor substrate, and N2 reduction is accompanied by considerable H2 evolution. Acetylene reduction leads to the production of some ethane as well as ethylene. Beyond this, little has been investigated. Under optimal conditions for N2 reduction, the ratio of N2 reduced to H2 produced was 1 7.5 compared with 1 1 for molybdenum nitrogenase 192). 

The development of the acetylene reduction assay to measure nitrogenase activity. 

This new assay technique was developed independently by American and Australian workers who first established the connection between nitrogenase and hydrogenase, and showed the former could reduce a number of substrates other than nitrogen, for example nitrous oxide, azide, cyanide and, most usefully, acetylene. The latter is reduced to ethylene and this forms the basis of the acetylene reduction test. The material to be examined is briefly gassed with acetylene and the ethylene formed measured by gas chromatography. While this technique was being refined,... 

Newell, S.Y., Hopkinson, C.S., and Scott, L.A., (1992) Patterns of nitrogenase activity (acetylene reduction) associated with standing, decaying shoots of Spartina altemiflora. Estuar. Coastal Shelf Sci. 35, 127-140. 

Capone, D. G. (1993). Determination of nitrogenase activity in aquatic samples using the acetylene reduction procedure. In Handbook of Methods in Aquatic Microbial Ecology (Kemp, P. F., Sherr, B. F., Sherr, E. B., and Cole, J. J., (eds.). Lewis Press, Boca Raton, FL. pp. 621—631. 

Patriquin, D. G., and McClung, C. R. (1978). In situ acetylene reduction assays of nitrogenase activity associated with the emergent halophyte Spartina alternijlora Loisel Methodological problems. Mar. Biol 47, 22-242. 

Ohki, K., and Fujita, Y. (1988). Aerohic nitrogenase activity measured as acetylene reduction in the marine non-heteerocystous cyanohacterium Trichodesmium spp. grown under artificial conditions. Mar. Biol. 91, 9-13. 

Nitrogenase catalyzes the reduction of many substrates (Table IV) and there are several lines of evidence to suggest that these are reduced at different sites or by different forms of the enzyme. Kinetic studies of interactions between pairs of substrates (Hwang ef al., J973 Rivera-Ortiz and Burris, 1975) show that Ng inhibits acetylene reduction competitively whereas the reverse inhibition is noncompetitive azide and acetylene are mutually noncompetitive and azide and cyanide inhibit each other competitively. HCN inhibits Nj reduction competitively whereas Nj inhibits HCN reduction in an undetermined manner. N2O inhibits Nj reduction competitively and acetylene reduction noncompetitively. ATP-dependent evolution by nitrogenase is inhibited by all the other reducible substrates and CO inhibits all substrate reductions except H2 evolution in a noncompetitive manner. Table V summarizes these results. The simplest interpretation of these results is that the N2-reducing site reduces N2 and possibly N2O but no other substrate azide, HCN, and methyl isocyanide are reduced at the same site... 

Acetylene, apparently, can be reduced by nitrogenase at high- and low-affinity sites either in vitro (Davis et al., 1975) or in vivo (Walker and Yates, 1978). Furthermore, preexposure to acetylene enhances the subsequent rate of acetylene reduction I /i vivo (David and Fay, 1977 David e/ al., 1978 Apte et al., 1978 Walker and Yates, 1978b) in the presence of chloramphenicol, which inhibits protein synthesis (Apte et al., 1978). It is likely, therefore, to be an acetylene-induced conformation change in the nitrogenase, but a possible effect on ATP synthesis or electron transfer to nitrogenase cannot be ruled out at this stage. 

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