Biomedical subjects

Yonghui Wang

Publications and source records attributed to Yonghui Wang.

23 records · Page 2Linked to original sources

PubMed1998

Radical Ion Probes. 9. The Chemistry of Radical Cations Derived from 9-Cyclopropylanthracene and 9-Bromo-10-cyclopropylanthracene.

Reactions of radical cations generated from 9-cyclopropylanthracene (1) and 9-bromo-10-cyclopropylanthracene (2) in the presence of methanol have been investigated electrochemically. The major products arising from oxidation of both substrates are attributable to CH(3)OH attack at the aromatic ring (occurring at the radical cation stage for 2 and the dication stage for 1) rather than CH(3)OH-induced cyclopropane ring opening, which is estimated to be exothermic by 20 kcal/mol. Although ring-opened products are detected in some instances, these are found to arise from subsequent reaction of the primary oxidation products. These observations are consistent with a proposed product-like transition state for the nucleophile-induced ring opening of cyclopropylarene radical cations in which the positive charge is localized on the cyclopropyl group and nucleophile, and thus unable to derive stabilization from the aromatic ring.

Journal Article↗

PubMedDate not supplied

Complexation with iron at sub-ppm levels in HPLC analyses of phospholipids.

Gradient RP-HPLC analysis of a phospholipid, E5564, utilizing water, methanol and phosphoric acid occasionally results in the appearance of a broad unknown peak in the chromatogram before a well-resolved E5564 peak. This unknown peak does not elute in a reproducible fashion with regards to peak shape and retention time, and is not present in chromatograms resulting from injections of the diluent alone. Investigation of this phenomenon revealed that iron ions in sub-ppm levels in the HPLC mobile phase chelated E5564 and the resultant complex(es) comprised the broad peak. The iron source was identified as phosphoric acid, which was used as a mobile phase modifier. Further studies were conducted to characterize the nature of the phospholipid-iron association and resultant complex.

Chromatography, High Pressure Liquid↗

PubMedDate not supplied

Proteomic profiling of Escherichia coli proteins under high cell density fed-batch cultivation with overexpression of phosphogluconolactonase.

In this study, we used proteomics to better understand the growth on glucose of Escherichia coli in high cell density, fed-batch cultures and the response to overexpression of plasmid-encoded 6-phosphogluconolactonase (PGL). Using liquid chromatography coupled to electrospray mass spectrometry, at least 300 proteins were identified in the cytosolic fraction of the six time points used to monitor the fermentation. The relative abundance changes of selected proteins were obtained by comparing the peak area of the corresponding peptides at a particular m/z (mass over charge ratio) value. During the time course of samples collected during the rapid growth achieved under batch and fed-batch conditions, both the control and recombinant E. coli strains showed up-regulation of proteins participating in the tricarboxylic acid (TCA) cycle, particularly acetyl-CoA synthetase (AcCoAS), malate dehydrogenase (MDH), and succinyl-CoA synthetase (SuccCoAS). In the recombinant strain culture, fumarase was up-regulated until 35 h after inoculation but was not in the control strain culture. In addition, the proteomic measurement detected up-regulation of three well-characterized binding transport proteins in both control and recombinant strains. The up-regulation of TCA cycle enzymes is consistent with the increase in growth rate observed in the cell culture. In addition, up-regulation of these proteins demonstrated the importance of both the pentose-phosphate shunt and TCA cycle to the increased biosynthetic activity required by a high level protein synthesis. This study shows the potential of proteomics using shotgun sequencing (LC/MS of tryptic digests) to measure global changes in protein abundance during a fermentation process and will facilitate the development of robust manufacturing systems.

Bioreactors↗

PubMedDate not supplied

Monitoring of glycoprotein products in cell culture lysates using lectin affinity chromatography and capillary HPLC coupled to electrospray linear ion trap-Fourier transform mass spectrometry (LTQ/FTMS).

In this paper, we describe the combination of lectin chromatography with capillary LC coupled to a linear ion trap-Fourier transform mass spectrometer (LTQ/FTMS) to enrich and characterize overexpressed glycoproteins from a cell culture lysate. A well-characterized glycoprotein, recombinant tissue plasminogen activator (rt-PA), was used as a standard, and we demonstrated that the three N-linked glycopeptides (including glycan structures) present in a tryptic digest of the rt-PA standard could be characterized in the new hybrid MS platform. A feature of this approach is that a significant amount of information can be obtained about the carbohydrate structures by direct analysis of the tryptic digest without the need for additional time-consuming sample preparation protocols. A combination of lectins was then studied for improved recovery of captured glycopeptides and was related to the selectivity of different lectins for specific glycosylation motifs. This approach was then extended to the lysate of a cell line routinely used in biotechnology manufacture (Chinese hamster ovary, CHO). This study showed that the combinations of lectins could enrich glycoproteins significantly from a CHO cell lysate. We also demonstrated that with this level of enrichment and with the new hybrid mass spectrometer, we could study the structures of N-linked glycopeptides of rt-PA present in a crude CHO cell lysate, at a ratio of 1:200 (rtPA:total cell lysate protein, w/w) by accurate mass measurement in the FTMS and tandem MSn in the linear ion trap. The generic and high throughput nature of the lectin approach combined with the ability to directly analyze the glycan structures in the tryptic digest suggest that this platform has the potential to routinely monitor glycoprotein products at early stage manufacturing in the biotech industry.

Animals↗

PubMedDate not supplied

A pharmacoproteomics study of the cancer cell line EKVX using capillary-LC/MS/MS.

The chemotherapeutic agent camptothecin, 10-OH (CPT,10-OH), was shown to act synergistically with the epithelial growth factor receptor (EGFR) inhibitor (AG1478) against several transformed cell lines. To study the cellular response to these drugs, the non-small-cell lung carcinoma cell line, EKVX, was treated with these compounds either alone or in combination. We performed a proteomic analysis using capillary-HPLC coupled with electrospray ion trap mass spectrometry (capillary-LC-ESI/MS) of a tryptic digest to obtain a global protein profile of the EKVX cell line and identify changes in protein expression. The combination of AG1478 and CPT,10-OH showed synergistic cytotoxicity and also changed the expression of multiple proteins, while individual treatments showed a lesser effect on protein expression. Thus, the synergistic action of AG1478 and CPT,10-OH was reflected in altered protein profiles, showing that a proteomic analysis can serve to evaluate chemotherapeutic agents and their combinations.

Amino Acid Sequence↗