Abstract
Publisher Summary This chapter provides an overview of a procedure for the evaluation of amino acid metabolism in maple syrup urine disease (MSUD).The MSUD is an autosomal recessive disease in which there is an inability to degrade the three branched-chain amino acids (BCAA)—leucine, isoleucine, and valine. In the method described in the chapter, one of the BCAA, which is labeled with radioactivity in the carboxyl carbon, is incubated with a suspension of fibroblasts. The CO2 is trapped in a center well and assayed for radioactivity. The BCAA is used as substrate, instead of the branched-chain keto acids (BCKA), because it provides a lower and more consistent blank. The first degradative step, transamination, does not appear to be rate limiting in the normal subject, and is certainly not rate limiting in MSUD. Suspensions of cells, instead of homogenates or cell extracts, are used because much of the enzyme activity is lost when the cells are broken. The amount of radioactive CO2 that is evolved is compared to the incorporation of the radioactive amino acid into protein. The ratio relates the decarboxylase activity to the amount of metabolically active tissue.
Published Version
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