Chapter 2 Evidence for Various 4-Vinyl Reductase Activities in Higher Plants

Summary

[4-Vinyl] Mg-protoporphyrin reductase, [4-vinyl] Mg-protoporphyrin monoester reductase, [4-vinyl] protochlorophyllide a reductase, and [4-vinyl] chlorophyllide a reductase activities were solubilized from isolated barley plastid membranes. Solubilization enhanced the 4-vinyl reductase activities. Preliminary comparative investigations were performed on [4-vinyl] protochlorophyllide a reductase and [4-vinyl] chlorophyllide a reductase. Both activities were localized in the inner plastid membranes and were missing from the plastid envelopes. Like [4-vinyl] chlorophyllide a reductase, [4-vinyl] proto­chlorophyllide a reductase exhibited an absolute requirement for NADPH, and both activities were insensitive to NADP and nicotinamide. However, [4-vinyl] protochlorophyllide a reductase activity appeared to be less sensitive to light treatments than [4-vinyl] chlorophyllide a reductase, since after 4 h of illumination the ratio of [4-vinyl] chlorophyllide a reductase/[4-vinyl] protochlorophyllide a reductase activities dropped from 67 to 43. This in turn suggested that [4-vinyl] chlorophyllide a reductase and [4-vinyl] protochlorophyllide a reductase activities may be catalyzed by two different enzymes. Also, the following information suggests that the various 4-vinyl reductase activities were catalyzed by different enzymes; (a) the different 4-vinyl reductase activities exhibited different activations following solubilization, and (b) the ratio of [4-vinyl] chlorophyllide reductase/[4-vinyl] protochlorophyllide a reductase activities was different in variously purified fractions. Contrary to previous beliefs, it became evident that [4-vinyl] chlorophyllide a reductase was as active at the end of the dark phase as in the middle of the light phase of the photoperiod, in dark-monovinyl-light divinyl-light-dark monovinyl plants such as barley, as well as in dark-divinyl-light divinyl-light-dark divinyl plants such as cucumber. The carboxylic Chl biosynthetic routes were revised to accommodate the above observations. As a consequence two different pathways comprising appropriate biosynthetic routes are presented for dark-monovinyl-light divinyl-light-dark monovinyl plants such as barley, and dark-divinyl-light divinyl-light-dark divinyl plants such as cucumber.

Note: Unless proceeded by MV or DV, tetrapyrroles are used generically to designate metabolic pools that may consist of MV and/or DV ­components. It has come to our attention that the various 4VCR enzymes may be coded for by a limited number of genes whose expression is modified by various regulatory factors that bind to a particular gene and modify its expression. Alternatively, a single mutation as in the Nec 2 corn mutant could disrupt multiple enzymes due to a mutation in a single regulatory factor (Wang et al., 1999; Rebeiz et al., 2010).