that activation of protein synthesis might be 

 controlled through the transient proteolytic 

 activity at fertilization. Here, the ribosomes are 

 visualized as being coated by a protein envelope, 

 thus preventing protein synthesis sterically. 

 They visualize this envelope as being removed 

 by the protease, thus resulting in increased 

 protein synthesis. 



The TPNH change could account for the 

 observed activation of lipid synthesis at fer- 

 tilization, since this coenzyme is specifically 

 involved in this synthetic sequence. The TPNH 

 change, especially the increase in the redox 

 couple of TPNH/TPN, could also be critical 

 for protein disulphide interactions believed to 

 be involved in cell division (38). The total 

 increase in the triphosphopyridine nucleotides 

 could also be involved in the channeling of 

 carbohydrate through the pentose shunt, whose 

 activity increases following fertilization (39, 

 40, 9). A change in carbohydrate flux, although 

 still not rigorously proven, could also be 

 important in regulating macromolecule synthe- 

 sis. Besides the important energy yields from 

 carbohydrate metabolism, a major limiting fac- 

 tor could be carbon skeletons for synthesis, as, 

 e.g., ribose for RNA synthesis. 



V. Conclusions 



The present study of the temporal sequence 

 and mechanism of the fertilization reactions in 

 sea urchin eggs has centered on light- scattering 

 (structural) changes, fertilization acid excre- 

 tion, and activation of DPN kinase and respira- 

 tion. The data indicate that changes in light- 

 scattering and acid excretion begin 

 simultaneously, followed almost immediately 

 by activation of DPN kinase. Respiratory ac- 

 tivity increases last. 



Analysis of these changes suggests that the 

 light- scattering and acid changes reflect the 

 breakdown of the cortical granules. DPN kinase 

 activation might be through the free Ca+2 re- 

 lease known to occur after fertilization since 

 this enzyme is both Ca+2 and Mg+2 activated. 

 The mechanism of respiratory activation is still 

 unclear, but the available data suggest substrate 

 mobilization, possibly through control of 

 glycogen phosphorylase. 



POLLARD: Is there any possibility of get- 

 ting at this genetically? Are there any deficient 

 eggs which require that a large amount of cal- 

 cium be added to the medium in order to get 



fertilization? This sort of thing would be some- 

 thing you could look at. There might be some- 

 thing here similar to the findings by Slonimski 

 on yeast mitochondria, which themselves are 

 rather specific kinetic things. Is this possible? 

 Are sea urchins accessible genetically? 



EPEL: Yes, generally they are. I think it 

 would be a very good contribution. There may 

 be some organisms in which you could do this. 

 You do require calcium to fertilize invertebrate 

 eggs in the sea water. 



POLLARD: I feel you're trying to describe 

 a lot of exciting kinetics without quite putting 

 your finger on the initiating point. 



EPEL: That's right. 



POLLARD: You think that the best lead so 

 far might very well be the potentiation of 

 enzyme by action of calcium or magnesium, 

 presumably initiated by some membrane com- 

 ponent that makes this possible. You're refer- 

 ring, essentially, to a fast physical change, 

 like chemiosmosis, followed by fairly rapid 

 concentration of an ion which is favorable to 

 enzyme X. I would feel that if you're starting 

 to look at a single enzyme, this is the sort of 

 thing that you could have missing genetically. 

 Then you'd have to add a whole lot of other 

 things to the medium to make it go. Is there 

 any evidence at all for this sort of thing? 



EPEL: Not that I know of. 



TS'O: Is this enzyme stimulated by pH 

 changes? For instance, will a simple change 

 of pH from 6.9 to 6.5 affect the enzyme activity? 



EPEL: No, it appears to have a broad 

 optimum between pH 7 and 8. 



TS'O: Can physical studies be made on 

 fragments of membrane? 



EPEL: There have been some enzyme 

 studies made on sea urchin egg cell cortexes. 

 They have a sodium-potassium-activated 

 ATPase. 



PAPACONSTANTINOU: This might impli- 

 cate a regulation between the hexose mono- 

 phosphate pathway and the Embden-Meyerhof 

 pathway of glycolysis. We know that some 

 substrates from the hexose-monophosphate 

 pathway will regulate the activity of some 

 glycolytic enzymes. I wonder whether there 

 might be some regulation here where sedo- 

 heptulose-7-phosphate or other metabolites of 

 this cycle aiffect the activity of this enzyme. 



EPEL: Yes, I think this would be very 

 possible. 



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