1. Predictingand testing enzymatic activity Predicting and testing enzymatic activity Charlie Harrison Supervisors: Profs. John Ward & Christine Orengo

2. Norcoclaurine Benzylisoquinoline alkaloids (BIQAs) – group of plant secondary metabolites Includes many interesting compounds Common entry point: norcoclaurine (S)-norcoclaurine(R)-norcoclaurine

3. Norcoclaurine NorcoClaurine Synthase – NCS Five known examples from three different species But BIQAs exist in many more species… Can we find more NCSs?

4. Main Project Aims Bioinformatics – Identify uncharacterised proteins that are likely to exhibit NCS activity Lab Work – Express these proteins in E.coli and test predictions

5. Bioinformatics Available data: sequence and structure Use these to predict function

6. Bioinformatics Reproduced from Orengo, Jones & Thornton 2003 with permission

7. Bioinformatics Structural data is better for identifying functional homologues...

8. Bioinformatics Structural data is better for identifying functional homologues...but is only available for ~1% of sequences Use predictions of structural regions – Gene3D Starting point: Gene3D family of likely structural relatives Sequence clustering

9. Bioinformatics

10. Lab Work Design and order the synthetic genes Wait 3 weeks…

11. Lab Work

12. Results ✓ ✓ ✗

14. Thanks