Substrate specificity of the short chain fatty acyl-coenzyme a synthetase of Pinus radiata

The short chain fatty acid-CoA synthetase of the female gametophytic tissue of Pinus radiata exhibited highest affinity for propionate and butyrate (K_m(app.) = 0.073 mM for both substrates) when examined by ³²PPi-ATP exchange. Relative to the activity at 1 mM propionate, partially purified enzyme supported exchange with acetate (100%), acrylate (114%), crotonate (107%) and thioglycollate (70%), but with varying degrees of affinity as reflected by their different K_m(app) values (2.56, 1.41,0.45 and 1.26 mM, respectively). All other unhalogenated C₂, C₃ and Q carboxylic acids examined did not support PPi-ATP exchange. Partially purified enzyme supported low rates of exchange in the presence of 2-chloroacetate, 2-bromoacetate, 3-chloropropionate, 3-bromopropionate, 2-chlorobutyrate and 2-bromobutyrate; the apparent K_m values were 2.74, 1.03, 0.15, 0.17, 0.85 and 0.23 mM, respectively. In the presence of propionate and one of the unhalogenated or halogenated substrates, the PPi-ATP exchange was consistent with competition for a common substrate binding site. All other halogenated derivatives of acetate and propionate did not support PPi-ATP exchange. The enzyme displayed an absolute requirement for Mg²⁺ (optimum concentration 1.5 mM).