Human secreted proteins play a very important role in signal transduction. In order to study all potential secreted proteins identified from the human genome sequence, systematic production of large amounts of biologically active secreted proteins is a prerequisite. We selected 25 novel genes as a trial case for establishing a reliable expression system to produce active human secreted proteins in Escherichia coli. Expression of proteins with or without signal peptides was examined and compared in E. coli strains. The results indicated that deletion of signal peptides, to a certain extent, can improve the expression of these proteins and their solubilities. More importantly, under expression conditions such as induction temperature, N-terminus fusion peptides need to be optimized in order to express adequate amounts of soluble proteins. These recombinant proteins were characterized as well-folded proteins. This system enables us to rapidly obtain soluble and highly purified human secreted proteins for further functional studies.

Imidazolonepropionase (EC 3.5.2.7) is the third enzyme of the histidine degradation pathway that has been conserved from bacteria to eukaryotes. The enzyme is the only one with unknown three-dimensional structure in this pathway. In this work, Bacillus subtilis imidazolonepropionase (HutI) was expressed in E. coli and purified to homogeneity. After thrombin digestion, high quality crystals were obtained by hanging-drop vapor diffusion method. The best crystal diffracted to 2.0 A° and belonged to the space group P21 with unit-cell parameters a=57.73A°, b=106.34A°, c=66.47A°, b=89.93.

The adrenal gland protein AD-004 like protein (ADLP) from Caenorhabditis elegans was cloned and expressed in Escherichia coli. Enzyme assays showed that ADLP has special adenylate kinase (AK) properties, with ATP and dATP as the preferred phosphate donors. In contrast to all other AK isoforms, AMP and dAMP were the preferred substrates of ADLP; CMP, TMP and shikimate acid were also good substrates. Subcellular localization studies showed a predominant nuclear localization for this protein, which is different from AK1–AK5, but similar to that of human AK6. These results suggest that ADLP is more likely a member of the AK6 family. Furthermore, RNAi experiments targeting ADLP were conducted and showed that RNAi treatment resulted in the suppression of worm growth.

The presence of intranuclear and cytoplasmic inclusions in cells of embryogenic callus and developing embryoids from tissue cultures of Panax quinque folius L. was described, These cellular structures were not found in non-embryogenic cells. The size of intranuclear and cytoplasmic inclusions seemed to be related to the developmental status of the cell.