D21B Isolated from Pollen Pots of the Australian Stingless Bee .

bacteria have been previously isolated predominantly from floral nectar and identified in metagenomic screenings as associated with bees. Here, we isolated three strains from the robust Australian stingless bee sharing over 99.4% sequence similarity with strains isolated from floral nectar.

Cytochrome P450cin (CYP176A1).

Cytochrome P450cin (P450cin) (CYP176A1) is a bacterial P450 enzyme that catalyses the enantiospecific hydroxylation of 1,8-cineole to (1R)-6β-hydroxycineole when reconstituted with its natural reduction-oxidation (redox) partner cindoxin, E. coli flavodoxin reductase, and NADPH as a source of electrons. This catalytic system has become a useful tool in the study of P450s as not only can large quantities of P450cin be prepared and rates of oxidation up to 1,500 min(-1) achieved, but it also displays a number of unusual characteristics.

The trivalent copper complex of a conjugated bis-dithiocarbazate Schiff base: stabilization of Cu in three different oxidation states.

The new tribasic N(2)S(2) ligand H(3)ttfasbz has been synthesized by condensation of 4-thenoyl 2,2,2-trifluoroacetone and S-benzyl dithiocarbazate. On complexation with copper(II) acetate, spontaneous oxidation to the Cu(III) oxidation state is observed, and the complex [Cu(ttfasbz)] has been isolated and characterized structurally. Reduction to the EPR active Cu(II) analogue has been achieved chemically and also electrochemically, and in both cases, the process is totally reversible.

Cytochrome P450(cin) (CYP176A1) D241N: investigating the role of the conserved acid in the active site of cytochrome P450s.

P450(cin) (CYP176A) is a rare bacterial P450 in that contains an asparagine (Asn242) instead of the conserved threonine that almost all other P450s possess that directs oxygen activation by the heme prosthetic group. However, P450(cin) does have the neighbouring, conserved acid (Asp241) that is thought to be involved indirectly in the protonation of the dioxygen and affect the lifetime of the ferric-peroxo species produced during oxygen activation. In this study, the P450(cin) D241N mutant has been produced and found to be analogous to the P450(cam) D251N mutant.

Oxygen activation by P450(cin): Protein and substrate mutagenesis.

A conserved threonine found in the majority of cytochromes P450 (P450s) has been implicated in the activation of dioxygen during the catalytic cycle. P450(cin) (CYP176A) has been found to be an exception to this paradigm, where the conserved threonine has been replaced with an asparagine. Prior studies with a P450(cin) N242A mutant established that the Asn-242 was not a functional replacement for the conserved threonine but was essential for the regio- and stereocontrol of the oxidation of cineole.