Citryl-CoA and the citrate condensing enzyme

Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/32278/1/0000340.pd

The citrate cleavage enzyme. III. Citryl coenzyme a as a substrate and the stereospecificlty of the enzyme

This article does not have an abstract

Rapid Diffusion of Green Fluorescent Protein in the Mitochondrial Matrix

Abstract. It is thought that the high protein density in the mitochondrial matrix results in severely restricted solute diffusion and metabolite channeling from one enzyme to another without free aqueous-phase diffusion. To test this hypothesis, we measured the diffusion of green fluorescent protein (GFP) expressed in the mitochondrial matrix of fibroblast, liver, skeletal muscle, and epithelial cell lines. Spot photobleaching of GFP with a 100× objective (0.8-μm spot diam) gave half-times for fluorescence recovery of 15–19 ms with >90% of the GFP mobile. As predicted for aqueous-phase diffusion in a confined compartment, fluorescence recovery was slowed or abolished by increased laser spot size or bleach time, and by paraformaldehyde fixation. Quantitative analysis of bleach data using a mathematical model of matrix diffusion gave GFP diffusion coefficients of 2–3 × 10−7 cm2/s, only three to fourfold less than that for GFP diffusion in water. In contrast, little recovery was found for bleaching of GFP in fusion with subunits of the fatty acid β-oxidation multienzyme complex that are normally present in the matrix. Measurement of the rotation of unconjugated GFP by time-resolved anisotropy gave a rotational correlation time of 23.3 ± 1 ns, similar to that of 20 ns for GFP rotation in water. A rapid rotational correlation time of 325 ps was also found for a small fluorescent probe (BCECF, ∼0.5 kD) in the matrix of isolated liver mitochondria. The rapid and unrestricted diffusion of solutes in the mitochondrial matrix suggests that metabolite channeling may not be required to overcome diffusive barriers. We propose that the clustering of matrix enzymes in membrane-associated complexes might serve to establish a relatively uncrowded aqueous space in which solutes can freely diffuse

Starch-gel electrophoresis of citrate-condensing enzyme from pig heart

Citrate-condensing enzyme from pig heart can exist in vitro as two distinct species which are separable by starch-gel electrophoresis. Several mild types of treatment can interconvert these enzymes and suggest that the separate forms arise in the process of purification; the two enzymes may differ only in the state of reduction of their sulfhydryl groups.Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/32276/1/0000338.pd

Incorporation of radioactive citrate into fatty acids

Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/32310/1/0000378.pd