Engineering of prenyltransferase NphB for improving the regioselective olivetolic acid and geranyl pyrophosphate
- 주제(키워드) Cannabinoids , Cannabigerolic acid , NphB prenyltransferase , rational design , enzyme engineering , FuncLib
- 주제(DDC) 547
- 발행기관 아주대학교 일반대학원
- 지도교수 Tae Hyeon Yoo
- 발행년도 2024
- 학위수여년월 2024. 8
- 학위명 석사
- 학과 및 전공 일반대학원 분자과학기술학과
- 실제URI http://www.dcollection.net/handler/ajou/000000034200
- 본문언어 영어
- 저작권 아주대학교 논문은 저작권에 의해 보호받습니다.
목차
1. Introduction 1
1.1. Cannabinoid 1
1.1.1. Cannabinoid derived drugs 3
1.1.2. Methods of producing cannabinoids 4
1.1.3. Biosynthetic pathway of cannabinoids 5
1.2. Prenylation of olivetolic acid (OA) and geranyl pyrophosphate (GPP) 6
1.3. Prenyltransferase NphB 7
1.3.1. Limitation of NphB on CBGA production 8
1.4. Aim of this study 9
2. Methods and Materials 10
2.1. Construction of plasmids 10
2.2. Expression and purification of proteins 21
2.3. Enzymatic assay 22
2.4. Detection of Products 23
2.5. Kinetic Assays 24
3. Results and discussion 25
3.1. Design of NphB mutants via FuncLib 25
3.2. Importance of S214 position for regioselective synthesis of CBGA 30
3.3. Docking OA in the NphB structure 33
3.4. Introducing mutation at Y288 36
3.5. Introducing further mutation on A232 38
3.6. Introducing previously reported important mutation V49W 41
3.7. Kinetic parameters of NphB mutants toward OA 43
4. Conclusion 45
5. References 46
