Eukaryotic expression: developments for structural proteomics

The production of sufficient quantities of protein is an essential prelude to a structure determination, but for many viral and human proteins this cannot be achieved using prokaryotic expression systems. Groups in the Structural Proteomics In Europe ( SPINE) consortium have developed and implemented high- throughput ( HTP) methodologies for cloning, expression screening and protein production in eukaryotic systems. Studies focused on three systems: yeast ( Pichia pastoris and Saccharomyces cerevisiae), baculovirusinfected insect cells and transient expression in mammalian cells. Suitable vectors for HTP cloning are described and results from their use in expression screening and protein-production pipelines are reported. Strategies for coexpression, selenomethionine labelling ( in all three eukaryotic systems) and control of glycosylation ( for secreted proteins in mammalian cells) are assessed.

Light-induced off-flavor development in cloudy apple juice

Cloudy apple juice has been found to develop off-flavors during storage in daylight. The development of off-flavors and volatile compounds was monitored in reconstituted juice prepared from 'Golden Delicious' and 'Fuji' apple concentrates stored in glass bottles under fluorescent light (3000 Ix, 8 degrees C). A strong metallic off-flavor was formed by photooxidation. A major contributor to the off-flavor was identified as 1-octen-3-one by gas chromatography-olfactometry. In addition, six volatile compounds, pentanal, 2-methyl-1-penten-3-one, hexanal, (E)-2-heptenal, 6-methyl-5-hepten-2-one, and (E)-2octenal, increased significantly after light exposure and could contribute to the off-flavor. Except for pentanal and hexanal, these volatiles were found only after light exposure. Higher levels of volatiles were observed in juice from 'Golden Delicious' apples than in juice from 'Fuji' apples, and this difference was consistent with higher levels of suspended solids. When the suspended solids were removed by centrifugation, the development of volatiles on exposure to light was reduced significantly.