EcoRI
| EcoRI | |||||||||
|---|---|---|---|---|---|---|---|---|---|
 Kristalografska struktura restrikcione endonukleaze EcoRI bez DNK | |||||||||
| Identifikatori | |||||||||
| Simbol | EcoRI | ||||||||
| Pfam | PF02963 | ||||||||
| InterPro | IPR004221 | ||||||||
| SCOP | 1na6 | ||||||||
| SUPERFAMILY | 1na6 | ||||||||
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EcoRI je endonukleazni enzim izolovan iz sojeva E. coli. On je deo restrikciono modifikacionog sistema.
EcoRI se koristi kao restrikcioni enzim u molekularnoj biologiji. On kreira lepljive krajeve sa prepustima na 5' krajevima. Sekvenca nukleinske kiseline gde enzim preseca je GAATTC, što je komplementarno sa sekvencom CTTAAG. Ovi krajevi su rotaciono simetrični.
Struktura
Primarna struktura
Poput mnogih restrikcionih endonukleaza, EcoRI sadrži PD..D/EXK motiv unutar svog aktivnog mesta. On se tipično koristi za izolaciji i restrikciju bakterijske plazmidne DNK. U EcoRI ovaj motiv se sastoji od ostataka P90, D91, E111, A112, K113(2)1.
Tercijarna i kvaternarna struktura
Ovaj enzim je homodimer podjedinica sa 31 kD, koje sadrže globularne domene sa α/β arhitekturom. Svaka podjedinica sadrži petlju izvan globularnog domena koja se obavija oko DNK pri vezivanju..[2][3]
EcoRI je kokristalisan sa sekvencom koju normalno preseca. Taj kristal je korišten za rešavanje strukture kompleksa 1QPS. Rešena kristalna struktura pokazuje da su podjedinice enzima homodimeri koji simetrično interaguju sa DNK.[2] U kompleksu, dva α-heliksa sa svake podjedinice zajedno formiraju heliksni svežanj.[4] Na interagujućim heliksima su ostaci Glu144 i Arg145, koji formiraju prsten putem kojeg dva aktivna mesta enzima komuniciraju.[5]
Reference
- ^ A. Grigorescu, M. Horvath, P. A. Wilkosz, K. Chandrasekhar and J. M. Rosenberg (2004). „The Integration of Recognition and Cleavage: X-Ray Structures of Pre-Transition State Complex, Post-Reactive Complex, and the DNA-Free Endonuclease”. Nucleic Acids and Molecular Biology. 14: 137—177. doi:10.1007/978-3-642-18851-0_6.[мртва веза]
- ^ а б Pingoud, A., Jeltsch, A. (2001). „Structure and function of type II restriction endonucleases”. Nucl. Acids Res. 29 (18): 3705—3727. PMC 55916
. PMID 11557805. doi:10.1093/nar/29.18.3705.
- ^ Kurpiewski, M. R., Engler, L. E., Wozniak, L. A., Kobylanska, A., Koziolkiewicz, M., Stec, W. J, Jen-Jacobson, L (2004). „Mechanisms of coupling between DNA recognition and catalysis in EcoRI endonucleases”. Structure. 12: 1775—1788. PMID 15458627. doi:10.1016/j.str.2004.07.016.
- ^ Bitinaite, J., D. A. Wah, Aggarwal, A. K., Schildkraut, I. (1998). „FokI dimerization is required for DNA cleavage”. Proc Natl Acad Sci U S A. 95 (18): 10570—5. PMC 27935 . PMID 9724744. doi:10.1073/pnas.95.18.10570.
- ^ Kim, Y. C., Grable, J. C., Love, R., Greene, P. J., Rosenberg, J. M. (1990). „Refinement of Eco RI endonuclease crystal structure: a revised protein chain tracing”. Science. 249 (4974): 1307—1309. PMID 2399465. doi:10.1126/science.2399465.
Literatura
- Nicholas C. Price; Lewis Stevens (1999). Fundamentals of Enzymology: The Cell and Molecular Biology of Catalytic Proteins (Third изд.). USA: Oxford University Press. ISBN 019850229X.
- Eric J. Toone (2006). Advances in Enzymology and Related Areas of Molecular Biology, Protein Evolution (Volume 75 изд.). Wiley-Interscience. ISBN 0471205036.
- Branden C; Tooze J.. Introduction to Protein Structure. New York, NY: Garland Publishing. ISBN 0-8153-2305-0.
- Irwin H. Segel. Enzyme Kinetics: Behavior and Analysis of Rapid Equilibrium and Steady-State Enzyme Systems (Book 44 изд.). Wiley Classics Library. ISBN 0471303097.
- William P. Jencks (1987). Catalysis in Chemistry and Enzymology. Dover Publications. ISBN 0486654605.
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