VARIOUS FORMS OF YEAST LACTATE 

 DEHYDROGENASE 



By A. P. Nygaard 

 Johan Throne Hoist's Institute for Nutrition Research, Blinder n, Oslo 



Several forms of lactate dehydrogenase have now been isolated from aerobic 

 bakers' yeast (Nygaard, 1958; 1959a, b). The method involved: (1) ex- 

 traction by the stirring of a yeast suspension with micro glass beads, (2) 

 fractionation with acetone, (3) adsorption on calcium phosphate gel, (4) 

 ammonium sulphate fractionation, and (5) separation on an N,N-diethyl- 

 aminoethylcellulose (DEAE-cellulose) column. The different forms 

 differed greatly in their affinity for DEAE-cellulose. In the following, three 

 fractions (lactate dehydrogenase (LDH) I, II and III) will be described; they 

 were eluted in that order from the column. 



As a reference, the homogeneous and crystalline LDH of Appleby and 

 Morton (1954, 1959) has been used. This enzyme is well defined and well 

 characterized and will be referred to as 'crystalline LDH'. The properties 

 have been described by Appleby and Morton, 1954, 1955, 1959a, b and 

 Appleby, 1957 (see Morton, 1958). 



COMPOSITION AND SPECTRAL PROPERTIES OF THE 

 ISOLATED ENZYMES 



Lactate Dehydrogenase I {LDH I) 



This was eluted with 0-04 m phosphate around pH 6-5. Flavin and haem 

 were eluted as one peak in the ratio 1:2. As in the crystalline enzyme of 

 Appleby and Morton (1954, 1959a, b), the a-band was located at 557 m/z 

 and the Soret band in the reduced and oxidized state was located at 423 m/* 

 and 413 m// respectively. In contrast to the crystalline enzyme, however, 

 the heights of the reduced and oxidized Soret band were about equal. The 

 extinction coefficient was about 100 X 10^ m~i cm^^ compared with 240 X 

 10' for the crystalline enzyme (Appleby and Morton, 1959a, b). Further- 

 more, all the reduced bands were broader for this fraction. A specific 

 proteolytic attack at the site involved in the binding of the haem may have 

 modified the spectrum. The enzyme gave a normal pyridine protohaemo- 

 chrome spectrum (Nygaard, 1959b). 



544 



