An Optimized Protocol for Bacterial Cell Disruption with Zirconium Beads and Recovery of Intracellular Tyrosine Phenol Lyase
The biotechnological production of intracellular tyrosine phenol lyase finds important applications in
pharmaceutical industry for the development of novel therapeutic molecules like L-DOPA. Cell disruption is one of the
important steps in the downstream processing of intracellular enzymes and bioprocess industry demands this operation to
be energy, cost and time efficient. The efficiency of the disruption process for the release of tyrosine phenol lyase from
Citrobacter freundii in a bead beater with zirconium beads was studied. Under optimum conditions of cell disruption
(0.1mm zirconium beads, 6mg/mL dcw cells, 1mM EDTA, 0.7mM DTT and 9 disruption cycle), the specific activity of
enzyme reached to 0.038 IU/mg. The stability of cell free tyrosine phenol lyase increased by 5.8-fold by addition of
EDTA and DTT. The disruption of C. freundii cells with beads seems to be effective as evidenced by transmission
electron microscopy. This cell free enzyme with enhanced stability can be used subsequently in suspension or
immobilized form for the biotransformtion reactions.
Keywords: Cell disruption, intracellular enzymes, tyrosine phenol lyase, Citrobacter freundii, bead beater, TEM.
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