Purification and Characterization of 3-Methylcrotonyl-Coenzyme A Carboxylase from Higher Plant Mitochondria

Substrate Specificity of the 3-Methylcrotonyl Coenzyme A (CoA) and Geranyl-CoA Carboxylases from Pseudomonas aeruginosa

“…S1 in the supplemental material). This is consistent with the behavior of P. citronellolis MCCase, which is specific for the MC-CoA substrate and has no detectable GCCase activity (8), as also found in pea leaf and potato mitochondria (2). A difference between the MCCases of P. aeruginosa and P. citronellolis is the apparent fivefold-higher affinity for the MCCoA substrate (K 0.5 s of 9.2 M and 43 M, respectively).…”

Induction of β‐methylcrotonyl‐coenzyme A carboxylase in higher plant cells during carbohydrate starvation: evidence for a role of MCCase in leucine catabolism

“…2A indicates that a major biotinylated polypeptide with an apparent molecular weight of 74000 was detected in sycamore mitochondria. It was assumed to be the biotinylated subunit of MCCase, as its apparent molecular weight was identical to that found in potato tubers [13]. In mitochondria from control cells a basal level of MCCase was observed, which remained constant ( Fig.…”

“…First, part of the activity of MCCase could h,~ve been lost during the preparation of the mitochondria. B owever, this hypothesis is unlikely since the techniques u, ed for the isolation and purification of the mitochondria h.tve previously been shown not to interfere with the MCCase a, tivity [13]. Second, we can hypothesize that many soluble p~ oteins accumulated in the matrix of the mitochondria of sl irved cells (in addition to MCCase).…”

“…The preparation of crude cell extracts, and the determination of MCCase activity were made according to the procedure of Alban et al [13], using NaH14CO3 as a substrate. The standard assay consisted of 50 mM Hepes (pH 8), 2.5 mM MgC12, 1 mM ATP, 2 mM DTT, 10 mM NaH14CO3 (37 MBq/mmol), 20 mM KC1, 0.4 mM [~-methylcrotonyl-CoA, and 20 ~tg of protein in a final volume of 200 /.tl.…”

“…Gerbling and Gerhardt [9] suggested that an extraperoxisomal pathway is involved in leucine catabolism, since they reported that peroxisomes are unable to carry out the biotin-dependent carboxylation of I]-methylcrotonyl-CoA. Interestingly plant MCCase has been purified and located in mitochondria [12,13]. MCCase, present in all plant organs, is usually the most prevalent of the biotin-containing enzymes [14], and its activity can be regulated by biotinylation of the apoenzyme [15].…”