A simple and cost-effective protocol for high-yield expression of deuterated and selectively isoleucine/leucine/valine methyl protonated proteins in Escherichia coli grown in shaker flasks
 

A simple and cost-effective protocol for high-yield expression of deuterated and selectively isoleucine/leucine/valine methyl protonated proteins in Escherichia coli grown in shaker flasks

Abstract

A simple and cost-effective protocol is presented for expression of perdeuterated, Ile/Leu/Val 1H/13C methyl protonated proteins from 100Â*ml cultures in M9â??++â??/D2O medium induced at high (OD600â??~â??10) cell density in shaker flasks. This protocol, which is an extension of our previous protocols for expression of 2H/15N/13C and 1H/13C labeled proteins, yields comparable quantities of protein from 100Â*ml cell culture to those obtained using a conventional 1 L culture with M9/D2O medium, while using three-fold less α-ketoisovaleric (1,2,3,4-13C4; 3,4â?²,4â?²,4â?²-d4) and α-ketobutyric (13C4; 3,3-d2) acid precursors.



Source: Journal of Biomolecular NMR