The carbamoyl region of the active site of aspartate transcarbamoylase from Escherichia coli was probed using an enzyme assay in which the two substrates were varied near their respective K_m 's. The inhibitors tested, some synthesized and some commercially available, were chosen to satisfy the structural requirements for binding to either the dicarboxylate or phosphate region with a substituent capable of extending into the carbamoyl region. However, the dicarboxylate based inhibitors were found to bind in an abnormal manner (unlike L-aspartate or succinate on which they were based). The carbamoyl region was found to contain a positively charged side-chain and preliminary results indicate that tetrahedral groups are not preferred over trigonal moieties. It is suggested that electrostatic stabilization of the negative charge which develops in the transition state may be a major factor in promoting catalysis. The identity of this charged group in the carbamoyl region is postulated to be His134 based on available X-ray diffraction data. The binding subsites of the active site of this enzyme were also found to be oriented in essentially the same plane. These results will greatly aid in the design of future mechanism-based
inhibitors to this enzyme that may have therapeutic value (at this time the mammalian enzyme is thought to have a similar catalytic mechanism). / Thesis / Master of Science (MS)
| Identifer | oai:union.ndltd.org:mcmaster.ca/oai:macsphere.mcmaster.ca:11375/23285 |
| Date | 03 1900 |
| Creators | Dennis, Paul |
| Contributors | Chan, W. W. -C., Biochemistry |
| Source Sets | McMaster University |
| Language | English |
| Detected Language | English |
| Type | Thesis |
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