Structure of 2-keto-3-deoxy-6-phosphogluconate aldolase at 2.8 Å resolution

The structure of 2-keto-3-deoxy-6-phosphogluconate aldolase has been extended to 2.8 Å resolution from 3.5 Å resolution by multiple isomorphous replacement methods using three heavy-atom derivatives and anomalous Bijvoet differences to 6 Å resolution (〈m〉 = 0.72). The replacement phases were improved and refined by electron density modification procedures coupled with inverse transform phase angle calculations. A Kendrew model of the molecule was built, which contained all 225 residues of a recently determined amino acid sequence, whereas only 173 were accounted for at 3.5 Å resolution. The missing residues were found to be part of the interior of the molecule and not simply an appendage. The molecule folds to form an eight-strand α/β-barrel structure strikingly similar to triosephosphate isomerase, the A-domain of pyruvate kinase and Taka amylase. With a knowledge of the sequence, the nature of the interfaces of the two kinds of crystallographic trimers have been examined, from which it was concluded that the choice of trimers selected in the 3.5 Å resolution work was probably correct for trimers in solution. The active site region has been established from the position of the Schiff base forming Lys144 but it has not been possible to confirm it conclusively in independent derivative experiments. An apparent anomaly exists in the location of Glu56 (about 25 Å from Lys144). The latter has been reported to assist in catalysis.     

Nucleotide and Deduced Amino Acid Sequences of a New Subtilisin from an Alkaliphilic <Emphasis Type="Italic">Bacillus</Emphasis> Isolate

The gene for a new subtilisin from the alkaliphilic Bacillus sp. KSM-LD1 was cloned and sequenced. The open reading frame of the gene encoded a 97 amino-acid prepro-peptide plus a 307 amino-acid mature enzyme that contained a possible catalytic triad of residues, Asp³², His⁶⁶, and Ser²²⁴. The deduced amino acid sequence of the mature enzyme (LD1) showed approximately 65% identity to those of subtilisins SprC and SprD from alkaliphilic Bacillus sp. LG12. The amino acid sequence identities of LD1 to those of previously reported true subtilisins and high-alkaline proteases were below 60%. LD1 was characteristically stable during incubation with surfactants and chemical oxidants. Interestingly, an oxidizable Met residue is located next to the catalytic Ser²²⁴ of the enzyme as in the cases of the oxidation-susceptible subtilisins reported to date.Received: 19 November 2002 / Accepted: 19 December 2002     

Helix-sense-selective polymerization of diphenyl-2-pyridylmethyl methacrylate with chiral anionic initiators

Polymerization of diphenyl-2-pyridylmethyl methacrylate was carried out with the complexes of organolithium compounds with 22 chiral ligands. Helix-sense-selectivity of the polymerization was largely affected by a slight structural difference of chiral ligands. (+)-(S)-2-(1-Pyrrolidinylmethyl)pyrrolidine was the most effective ligand in producing a one-handed helical polymer with narrow molecular weight distribution.