Proteomic analysis of proteins differentially expressed in conidia and mycelium of the entomopathogenic fungus Aschersonia placenta.

The infection of insects by the entomopathogenic fungus Aschersonia placenta depends on conidia. To identify proteins differentially expressed in A. placenta conidia vs mycelia, we performed a comparative proteomic analysis of A. placenta using 2-dimensional gel electrophoresis (2-DE) and matrix-assisted laser desorption ionization/time-of-flight mass spectrometry (MALDI-TOF-MS). We detected 1022 2-DE protein spots in conidia and 1049 in mycelia and analyzed 48 (13 from conidia and 35 from mycelia) using MALDI-TOF-MS. Finally, we identified 28 proteins (7 from conidia and 21 from mycelia). The identified proteins exclusive to conidia included major proteins participating in oxidation-reduction processes and vegetative insecticidal protein 1 (Vip1), a protein that is likely involved in pathogenicity. The identified proteins exclusive to mycelia were those involved in biosynthesis and metabolism, including uridine diphosphate galactopyranose mutase, which might play key roles in hyphal morphogenesis. This report provides the first proteomic analysis of different developmental stages of an Aschersonia species. Although only a small number of proteins were identified, the data represent a useful foundation for future studies concerning the molecular basis of entomopathogenicity in the species A. placenta and in the genus Aschersonia.

Stereospecific synthesis of alkylidenecyclopropanes via sequential cyclopropene carbomagnesation/1,3-carbon shift.

Alkylidenecyclopropanes can be synthesized from enantiomerically enriched cyclopropene derivatives with >99% stereotransfer and good to excellent yield. The protocol comprises the stereoselective reaction of Grignard reagents with 1-alkoxymethyl-3-hydroxymethylcyclopropenes and a stereospecific [1,3] carbon shift reaction.

Octameric structure of the human bifunctional enzyme PAICS in purine biosynthesis.

Phosphoribosylaminoimidazole carboxylase/phosphoribosylaminoimidazole succinocarboxamide synthetase (PAICS) is an important bifunctional enzyme in de novo purine biosynthesis in vertebrate with both 5-aminoimidazole ribonucleotide carboxylase (AIRc) and 4-(N-succinylcarboxamide)-5-aminoimidazole ribonucleotide synthetase (SAICARs) activities. It becomes an attractive target for rational anticancer drug design, since rapidly dividing cancer cells rely heavily on the purine de novo pathway for synthesis of adenine and guanine, whereas normal cells favor the salvage pathway. Here, we report the crystal structure of human PAICS, the first in the entire PAICS family, at 2.8 A resolution. It revealed that eight PAICS subunits, each composed of distinct AIRc and SAICARs domains, assemble a compact homo-octamer with an octameric-carboxylase core and four symmetric periphery dimers formed by synthetase domains. Based on structural comparison and functional complementation analyses, the active sites of SAICARs and AIRc were identified, including a putative substrate CO(2)-binding site. Furthermore, four symmetry-related, separate tunnel systems in the PAICS octamer were found that connect the active sites of AIRc and SAICARs. This study illustrated the octameric nature of the bifunctional enzyme. Each carboxylase active site is formed by structural elements from three AIRc domains, demonstrating that the octamer structure is essential for the carboxylation activity. Furthermore, the existence of the tunnel system implies a mechanism of intermediate channeling and suggests that the quaternary structure arrangement is crucial for effectively executing the sequential reactions. In addition, this study provides essential structural information for designing PAICS-specific inhibitors for use in cancer chemotherapy.

Selective syntheses of Δ(α,ß) and Δ(ß,γ) butenolides from allylic cyclopropenecarboxylates via tandem ring expansion/[3,3]-sigmatropic rearrangements.

Allylic cyclopropenecarboxylates undergo ring expansion reactions to give 2-allyloxyfuran intermediates, which subsequently rearrange to Δ(ß,γ) butenolides via a Claisen rearrangement or to the corresponding Δ(α,ß) butenolides via further Cope rearrangement. Also described are methods for chirality transfer in the rearrangement of nonracemic allylic esters.