Oligomeric Forms of Pyruvate Kinase from Chlorella with Different Kinetic Properties

Fast protein liquid chromatography on Superose 6 o f crude extracts from the chlorophyllfree mutant no. 20 o f the unicellular green alga Chlorella kessleri reveals two possibly o ligo­ meric forms o f pyruvate kinase (2.7.1.40). Their occurrence is markedly altered in the course o f heterotrophic growth with changing levels o f exogenous glucose as carbon source with only one enzyme species with a MW o f 400 kDa existing in growing cells, two forms o f 400 and 580 kDa in resting cells. Substrate affinity towards PEP of the 400 kDa form is better than that o f the 580 kDa species; responses to the effector AM P are different as well. In vitro, addition o f PEP or o f AMP leads to the formation o f higher MW enzyme species with MW o f 730, 1050 and 1400 kDA without affecting the total activity. In vivo alterations in the levels o f several metabolites including PEP upon addition o f glucose have been shown to occur. Therefore, it is discussed, whether changes in the concentration o f intermediates and effectors may provide the mechanism for the increased rate o f carbohydrate degradation by affecting the occurrence and/or ratio o f the various PK forms with different kinetic and regulatory properties. Upon blue light irradiation, which also stimulates carbohydrate breakdown o f the Chlorella mutant cells, the distribution o f PK is shifted towards the species with higher substrate affinity, a re­ sult being in accordance with the above conception.