Table III.

Catabolism of Leu by mitochondrial extracts

Additions/OmissionsIncubation Durationα-KICIV-CoAMC-CoA
h dpm (×103)/mg protein
None0.33256.4  (+45.1)ND2-a ND
1585.3  (+102.9)19.7  (+2.3)ND
21303.2  (+229.1)110.5  (+13.0)ND
4697.6  (+122.7)426.5  (+50.2)8.6  (+4.2)
6362.1  (+63.7)508.6  (+59.8)29.9  (+14.4)
−α-KG289.4  (+15.7)NDND
−CoA21214.7  (+213.6)NDND
+1 mm α-KIC21099.0  (+193.2)NDND
41835.0  (+322.6)NDND
62270.3  (+399.1)NDND
+Avidin2936.8  (+164.7)154.3  (+18.2)ND
4800.7  (+139.9)420.6  (+49.5)51.4  (+24.7)
6602.4  (+105.9)436.8  (+51.4)71.7  (+34.5)

Mitochondrial extracts were incubated with [U-14C]Leu and cofactors for various lengths of time and the resulting radioactive products were analyzed by HPLC. Additional reactions were also conducted without added α-ketoglutarate (α-KG) or CoA, or with the addition of 1 mm nonradioactive α-KIC or 10 μg of avidin. α-KG is required for the transamination of Leu; CoA is required for the decarboxylation and activation of α-KIC in the BCKDH reaction; nonradioactive α-KIC inhibits further metabolism of α-[14C]KIC by competition; and avidin inhibits values are reported as in Table II.

    • F2-a ND, Not detected.