Project Details
Projekt
Correlation of structure and properties of protein crystals in consideration of downstream processing and formulation
Applicants
Dr. Rebekka Biedendieck; Professorin Dr. Anett Schallmey; Professor Dr.-Ing. Carsten Schilde
Subject Area
Biological Process Engineering
Mechanical Process Engineering
Mechanical Process Engineering
Term
from 2016 to 2024
Project identifier
Deutsche Forschungsgemeinschaft (DFG) - Project number 315462951
Because of their high volumetric catalytic activity and their high chemical and thermal resistance, enzymes in protein crystals are an excellent choice for application as immobilized biocatalysts. Since enzyme activity, product release rate as well as mechanical rigidity are dependent on crystal shape and size, their molecular basis shall be investigated. For this, the complete process chain consisting of the genetic modification, production, purification, crystallization, cross-linking and mechanical characterization of the model proteins halohydrin dehalogenase HheG and Penicillin G acylase PGA has been established in the first funding period. Three new structures of PGAs exhibiting different thermostabilities have been solved. For both model proteins, many genetic variants have been generated, successfully produced and tested towards their activity and thermal stability. The most promising candidates have been selected for the generation of native and cross-linked protein crystals in order to investigate their mechanical and catalytic properties. Currently, the particle structure of the protein crystals is modelled by Discrete Element Method to emulate their properties. Main focus of the second funding period will be the systematic modification of the proteins by genetic and chemical methods for an efficient cross-linking of the resulting protein crystals. In this respect, HheG will be used for a detailed investigation of the impact of cross-linking on the mechanical rigidity of the crystals, whereas the different PGAs will be used to test the transfer of gained knowledge from one to another protein. Next to this systematic engineering of enzymes, the scale up of the whole process chain will be established in order to determine the mechanical behavior of cross-linked protein crystals in model processes. Additionally, statistical and numerical methods will be used to explain observed changes of mechanical behavior based on protein interactions within the crystal as well as cross-linking sites. In summary, structural principles of crystal formation and cross-linking will be elucidated with the aim to obtain thermally and mechanically stable protein crystals for biotechnological application based on systematic genetic modification of the proteins.
DFG Programme
Priority Programmes
Co-Investigator
Dr.-Ing. Ingo Kampen
