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Using stable-isotope tracers, we recently confirmed that obesity-associated glycine deficiency compromised the synthesis and clearance of endogenous and xenobiotic acylCoAs as acylglycines 3

P., Bindschedler, L

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50 L of tissue homogenate was added to 750 L of phosphate buffer (0.1 mM, pH 7.5)

Furthermore, purified CFE systems by design contain no chaperones and extract-based systems may or may not have sufficient levels of endogenous chaperones to adequately support proper folding, and thus CFE systems are frequently supplemented with additional chaperones (e.g., DnaK/DnaJ/GrpE, GroEL/GroES)