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Human apolipoprotein D (ApoD) occurs in plasma associated with high density lipoprotein. Apart from the involvement in lipid metabolism, its binding activity for progesterone and arachidonic acid plays a role in cancer development and neurological diseases. The crystal structures of free ApoD and its complex with progesterone were determined at 1.8A resolution and reveal a lipocalin fold. The narrow, mainly uncharged pocket within the typical beta-barrel accommodates progesterone with its acetyl side chain oriented toward the bottom. The cavity adopts essentially the same shape in the absence of progesterone and allows complexation of arachidonic acid as another cognate ligand. Three of the four extended loops at the open end of the beta-barrel expose hydrophobic side chains, which is an unusual feature for lipocalins and probably effects association with the high density lipoprotein particle by mediating insertion into the lipid phase. This mechanism is in line with an unpaired Cys residue in the same surface region that can form a disulfide cross-link with apolipoprotein A-II.

Human apolipoprotein D (ApoD) is a physiologically important member of the lipocalin protein family that was discovered as a peripheral subunit of the high-density lipoprotein (HDL) but is also abundant in other body fluids and organs, including neuronal tissue. Although it has been possible to produce functional ApoD in the periplasm of Escherichia coli and to demonstrate its ligand-binding activity for progesterone and arachidonic acid, the recombinant protein suffers from a pronounced tendency to aggregate and to adsorb to vessel surfaces as well as chromatography matrices, thus hampering further structural investigation. Here, we describe a systematic mutagenesis study directed at presumably exposed hydrophobic side chains of the unglycosylated recombinant protein. As a result, one ApoD mutant with just three new amino acid substitutions--W99H, I118S, and L120S--was identified, which exhibits the following features: (1) improved yield upon periplasmic biosynthesis in E. coli, (2) elution as a monomeric protein from a gel permeation chromatography column, and (3) unchanged binding activity for its physiological ligands. In addition, the engineered ApoD was successfully crystallized (space group I4 with unit cell parameters a = 75.1 A, b = 75.1 A, c = 166.0 A, alpha = beta = gamma = 90 degrees), thus demonstrating its conformationally homogeneous behavior and providing a basis for the future X-ray structural analysis of this functionally still puzzling protein.

A highly substrate-specific microsomal-bound cytochrome P-450 NADPH and O2-dependent enzyme system has been discovered in immature seeds of Colchicum autumnale L. that is responsible for the intramolecular phenol-oxidative coupling of autumnaline to isoandrocymbine during colchicine biosynthesis.

The roots of Delphinium denudatum have yielded a new diterpenoid alkaloid, 8-acetylheterophyllisine (1), in addition to the known alkaloids vilmorrianone (2), panicutine (3), denudatine (4), isotalatizidine (5), and condelphine (6), as well as 3-hydroxy-2-methyl-4H-pyran-4-one (7). Compounds 1, 3, and 7 have not been isolated from this plant previously. The structures of compounds 1, 3, and 7 were determined through spectral and X-ray diffraction analyses. Compounds 1-3 have shown antifungal activity against a number of human pathogenic fungi.