529-0739-10L Biological Chemistry A: Technologies for Directed Evolution of Enzymes
|Autumn Semester 2018
|P. A. Kast, D. Hilvert
|yearly recurring course
|Language of instruction
|Advanced laboratory course or internship depending on lab course Biological Chemistry B
Candidates must inquire with P. Kast no later than September 1st whether course will take place (no self-enrollment)
Further information to registration and work hours: Link
|During this semester course, methodologies will be taught for biological-chemical enzyme evolution experiments using molecular genetic mutation technologies and in vivo selection in recombinant bacterial strains.
|All technologies used for the experiments will be explained to the students in practice with the goal that they will be able to independently apply them for the course project and in future research endeavors. After the course, an individual report about the results obtained has to be prepared.
|This class conducts and supports experiments for a specifically designed genuine research project. We will carry out biological-chemical enzyme evolution experiments using molecular genetic mutation technologies and in vivo selection in recombinant bacterial strains. The relevant technologies will be taught to the students, such as the preparation of competent cells, production and isolation of DNA fragments, transformation of gene libraries, and DNA sequencing. The course participants will generate a variety of different variants of a chorismate mutase. Individual enzyme catalysts will be purified and subsequently characterized using several different spectroscopic methods. The detailed chemical-physical analyses include determination of the enzymes' kinetic parameters, their molecular mass, and the integrity of the protein structure. The students will present the results obtained from their individual evolution experiments at the end of the semester. We expect that during this lab course we will not only generate novel enzymes, but also gain new mechanistic insights into the investigated catalyst.
|The necessary documents and protocols will be distributed to the participants during the course.
|General literature to "Directed Evolution" and chorismate mutases, e.g.:
– Taylor, S. V., P. Kast & D. Hilvert. 2001. Investigating and engineering enzymes by genetic selection. Angew. Chem. Int. Ed. 40: 3310-3335.
– Jäckel, C., P. Kast & D. Hilvert. 2008. Protein design by directed evolution. Annu. Rev. Biophys. 37: 153-173.
– Roderer, K. & P. Kast. 2009. Evolutionary cycles for pericyclic reactions – Or why we keep mutating mutases. Chimia 63: 313-317.
Further literature will be indicated in the distributed script.
|Prerequisites / Notice
|- This laboratory course will involve experiments that require a tight schedule and (sometimes) long (!) working days.
- The projects of this course are tightly linked to the ones of the Biology BSc course "529-0739-01 Biological Chemistry B: New Enzymes from Directed Evolution Experiments", which takes place as a block course during the month of November. There will be joint lectures for the participants of both courses during that time. The teaching language is English.
- The number of participants for the laboratory class is limited. It is mandatory to sign up for the course directly with P. Kast no later than September 1, prior to the start of the fall semester. Until then it will be decided whether the course will take place.
- A valid registration is considered a commitment for attendance of the entire semester course, as involved material orders and experimental preparations are necessary and, once the class has started, the flow of the experiments must not be interrupted by individual absences. In case of an emergency, please immediately notify P. Kast.
- For more information, see also Link or contact P. Kast directly (HCI F 333, Tel. 044 632 29 08, Link).