32 ALBERT L. LEHNINGER 



Respiratory chain phosphorylation, the most prominent mitochondrial 

 activity, is not only a very complex enzyme system, but one which differs 

 from many other enzyme systems in that it may never be understood in 

 mechanism until the active sites of the individual enzymes are identified, 

 since there are apparently no low-molecular weight diffusible inter- 

 mediates. The active sites of the enzymes involved are thus the "inter- 

 mediates". No amount of indirect experimentation or descriptive, physio- 

 logical study of the mitochondrion can thus replace direct isolation and 

 chemical study of the catalysts of energy-coupling and the catalysts activa- 

 ting the swelling-contraction cycle. 



The three most conspicuous properties of mitochondria are {a) the 

 catalysis of respiration and energy coupling, {b) the occurrence of reversible 

 swelling and contraction, leading to water movements, which are geared 

 to respiration and (r) ion transport, also geared to the respiratory chain. 

 After considering the mitochondrial membranes, in which these functions 

 apparently reside, this paper will deal with recent work on some isolated 

 components involved in these functions. 



Molecular organization of the mitochondrial membranes 



It is now clear from many items of evidence that the enzymes of 

 respiration and coupled phosphorylation are more or less firmly embedded 

 in or on the mitochondrial membranes ; indeed circumstantial evidence 

 suggests that the inner membrane which presumably forms the cristae 

 is the site of these enzymic activities. When the mitochondria are 

 subjected to disruption by either digitonin or sonic oscillation, they shatter 

 into fragments having a wide spectrum of particle weights. We have 

 examined the enzymic properties of a series of such fragments differing 

 in sedimentation rate and have found that they have a fairly constant 

 content per mg. protein N of cytochrome oxidase, ^-hydroxybutyric 

 dehydrogenase, succinoxidase, and ATP-ase, regardless of particle size 

 [2], suggesting that the membranes are made up of a large number of 

 recurring structural units, each of which may contain a complete assembly 

 of respiratory carriers in finite ratio, as determined by difference spectra. 



Calculations suggest that an individual liver mitochondrion may 

 contain 5000-10 000 or more, of such respiratory assemblies, which are 

 more or less evenly distributed on the membrane. Extension of such 

 calculations, with certain assumptions, indicates that a large fraction of 

 the total mass of the membrane is made up of these assemblies of catalyti- 

 cally active molecules — perhaps as much as 40% by weight [2]. The 

 membranes also contain considerable " phosphoprotein " and the phosphate 

 groups undergo replacement at a high rate. The exact disposition of 

 protein and lipid molecules in the membranes is not yet clear. The original 



