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5.3.2.1 Preparation of a crude extract
The extract will contain a complex mixture of all the proteins from the cell cytoplasm, and some additional macromolecules, cofactors and nutrients. Crude protein extracts are prepared by removal of cellular debris generated by cell lysis, which is achieved using chemicals and enzymes, sonication or a French Press. The debris are removed by centrifugation and the supernatant is recovered. Crude preparations of extracellular proteins may be obtained by simply removing the cells by centrifugation. Removal of denatured proteinsAn easy approach to purifying a heat-resistant protein is to denature the other proteins in the mixture by heating and then cooling the solution (thus allowing the thermo stable enzyme to reform or redissolve, if necessary). The denatured proteins can then be removed by centrifugation. /font> |