134 ALEXANDER L. DOUNCE 



fact that the enzyme was found by Lan"^ in high concentrations in Uver 

 cell nuclei isolated at pH 6.0 in very dilute citric acid. Again, D-amino oxi- 

 dase is an insoluble mitochondrial enzyme and thus will undoubtedly be 

 present in the fine mitochondrial fragments produced by the action of the 

 Waring Blendor when it is run rapidly enough to disrupt a high proportion 

 of liver cells. No D-amino oxidase was found in cell nuclei isolated by Lang 

 and Siebert.i^o 



Before consideration is given to other classes of nuclear enzymes, most of 

 which are water-soluble, the reader should be reminded that what has been 

 said concerning methods of estimating soluble proteins of the cell nucleus 

 applies for the most part to determinations of nuclear enzymes. It is still a 

 matter of dispute as to whether the nuclear membrane is permeable to 

 water-soluble proteins or not, but Andersoni2i-i23 ^^s assembled a fair amount 

 of evidence in favor of permeability, against which there is only rather 

 meager evidence to the contrary.^ •^■"** In any case, it can be shown by 

 direct experiment that under experimental conditions frequently employed 

 in isolating nuclei, protein can be extracted.* ■*'•''* Accordingly, one must 

 always anticipate the possible loss of a given soluble nuclear enzyme during 

 the isolation procedure. Such a loss for instance has apparently been directly 

 demonstrated by Mirsky for nucleoside phosphorylase.^* The use of a high 

 pH or a very low pH or saline in the homogenizing medium can be expected 

 to produce a particularly severe loss in soluble nuclear enzymes.*' 



Judging from work with arginase,* soluble enzymes are not necessarily 

 adsorbed to an appreciable extent from the aqueous homogenate by the 

 nuclei. Such adsorption of adenosinetriphosphatase by nuclei may occur, 

 according to results obtained by Mirsky et al.,"^^ although this might pos- 

 sibly be another instance of adsorption of finely divided mitochondrial 

 fragments carrying the adenosinetriphosphatase rather than an adsorption 

 of a soluble enzyme. 



In general, different methods for isolating cell nuclei in aqueous media 

 seem to lead to more concordant results for water-soluble, than for water- 

 insoluble, enzymes, although even here not all results are in agreement. 



Owing to the possibility of loss of a given enzyme during the extraction 

 procedure, on the one hand, or adsorption of enzymes by the nuclei, on 

 the other hand, Mirsky et alP maintain that the Behrens type of procedure 

 should always be used for a study of nuclear enzymes when this type of 

 isolation does not cause serious damage to the enzymes being studied. 

 Unfortunately, these authors do not list the extent of damage to the en- 

 zymes studied by them, although, owing to possible differences in degree of 



»2o K. Lang and G. Siebert, Biochem. Z. 320, 402 (1950). 

 '" N. G. Anderson, Science 117, 517 (1953). 

 1" N. G. Anderson, Exptl. Cell. Research 4, 306 (1953). 

 »" N. G. Anderson, J. Tenn. Acad. Set. 27, 198 (1952). 



