Expression systems, heme

The success of Chapman and co-workers in expression of flavocytochrome 2 in E. coli [23] is encouraging in its impUcations for future expression of flavoproteins in this host because, in their experience both the flavin and heme groups are incorporated into the recombinant protein. Moreover, the bacterial expression system produces the protein 500-1000 fold more efficiently than the yeast from which it was cloned. The enzyme produced in E. coli, however, lacks the first five amino acid residues at its amino terminus, a result which presumably reflects subtle differences in protein synthesis between the two organisms. 

The widespread expression of biliverdin reductase, which parallels the widespread expression of heme oxygenase, suggests that bilirubin may play a special role in particular tissues. Studies suggested that bilirubin is an excellent lipid-soluble antioxidant. Bilirubin reacts with reactive oxygen species (ROS) that oxidize bilirubin back to biliverdin. The biliverdin then is reduced again to bilirubin. This repeated cycling results in the net use of NADPH to protect cells from ROS (Fig. 22-1). This system may be especially important in neurons. 

One of the issues in P450 2E1 reactions is the need for b, first demonstrated with the rat enzyme and also the human enzyme - the involvement also exists in microsomes. also augments P450 2E1 activity in bacterial expression systems -. In contrast to several of the P450s, apo-fej (minus heme) does not function, arguing for a classic role of electron donation in enhancement of catalysts -. ... 

Nevertheless, the copurification of HOS and HAS observed in bacterial systems support the notion that HOS and HAS form a physiologically relevant complex. Furthermore, the fact that the presence of HAS appears to decrease the activity of HOS is consistent with the hypothesis that heme O is transferred directly from HOS to HAS, and that either the conversion of heme O into heme A or the release of the heme A product is the rate-determining step in this heterologous expression system. Thus, the heme A biosynthetic pathway may be at least partially regulated by product release. 

The heme dioxygenase enzymes involved in tryptophan oxidation catalyse the first and rate-limiting step in the kynurenine pathway—the 02-dependent oxidation of 1-tryptophan to AT -formylkynurenine. In the past 10 years, there have been substantial new developments, including new structural information, bacterial expression systems for a number of dioxygenases, contributions from computational chemistry, and emerging... 

In the discussion of the Kramer s doublets we have dealt entirely with the properties of the atomic orbitals of the heme iron. In a different approach the electronic states are expressed in terms of molecular orbitals. This is particularly interesting for aromatic radicals because the unpaired electron spreads out over the entire conjugated system of -orbitals. 

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