TY - JOUR
T1 - [79] Resolution and Reconstitution of ATP Synthesis and ATP-Dependent Functions of Liver Mitochondria
AU - Pedersen, Peter L.
AU - Hullihen, Joanne
N1 - Funding Information:
This work was supported by N1H Grant CA 10951 from the National Cancer Institute.
PY - 1979/1/1
Y1 - 1979/1/1
N2 - The concentration of urea is critical in preparing urea particles that are reconstitutively active in catalyzing adenosine triphosphate (ATP) synthesis. If 8 M urea is used to treat inner membrane vesicles in place of the 6 or 6.4 M urea, 4 M urea particles are obtained. These particles, in analogy to 3 or 3.2 M urea particles, readily bind purified F1 ATPase. However, the Fl-restored, 4 M urea particles have <30% maximal capacity to catalyze adenosine triphosphate (ATP) synthesis. F1-restored, 3 or 3.2 M urea particles, on the other hand, catalyze rates of ATP synthesis that are comparable to rates catalyzed by the starting inner membrane fraction. It is also critical for best results to assay the F1-restored urea particles immediately after centrifugation and resuspension. Failure to do so within 30 min to 1 hr results in loss of the ATP or AMP-PNP activating effect and suboptimal rates of ATP synthesis is observed.
AB - The concentration of urea is critical in preparing urea particles that are reconstitutively active in catalyzing adenosine triphosphate (ATP) synthesis. If 8 M urea is used to treat inner membrane vesicles in place of the 6 or 6.4 M urea, 4 M urea particles are obtained. These particles, in analogy to 3 or 3.2 M urea particles, readily bind purified F1 ATPase. However, the Fl-restored, 4 M urea particles have <30% maximal capacity to catalyze adenosine triphosphate (ATP) synthesis. F1-restored, 3 or 3.2 M urea particles, on the other hand, catalyze rates of ATP synthesis that are comparable to rates catalyzed by the starting inner membrane fraction. It is also critical for best results to assay the F1-restored urea particles immediately after centrifugation and resuspension. Failure to do so within 30 min to 1 hr results in loss of the ATP or AMP-PNP activating effect and suboptimal rates of ATP synthesis is observed.
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U2 - 10.1016/0076-6879(79)55081-2
DO - 10.1016/0076-6879(79)55081-2
M3 - Article
C2 - 156857
AN - SCOPUS:0018370087
SN - 0076-6879
VL - 55
SP - 736
EP - 741
JO - Methods in enzymology
JF - Methods in enzymology
IS - C
ER -