| Budget Amount *help |
¥3,800,000 (Direct Cost: ¥3,800,000)
Fiscal Year 2004: ¥1,100,000 (Direct Cost: ¥1,100,000)
Fiscal Year 2003: ¥2,700,000 (Direct Cost: ¥2,700,000)
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| Research Abstract |
It is well known that pyruvate kinase M1 and M2 isozymes are expressed from the single gene locus, PK-M gene. While the M2 isozyme undergoes allosteric regulation with its own substrate, phosphoenolpyruvate, and fructose-1,6-bisphosphate(FBP), the M1 is a non-allosteric isozyme, the activity of which is not altered by such allosteric effectors. However, we have found that the M1 isozyme exhibits cooperative response toward the change in Mg^<2+> whereas the M2 does not. In this study, we examined the kinetic properties of these enzymes in terms of the activity regulation by the divalent cation. In the case of M1,the sigmoid response with the increasing concentration of Mg^<2+> was observed while the response of M2 followed a simple saturation kinetics. Therefore, it was found that the M1 is uniquely regulated by a possible conformational change associated with Mg ion although it has been believed that the non-allosteric isozyme does not undergo allosteric regulation because of its persistent active conformation. These results and other findings suggest that the non-allosteric pyruvate kinase isozyme is regulated by a cooperative interaction between the subunits although such alteration is not caused by usual allosteric effectors. Replacement of Tyr-389 in the M2 by the corresponding residue in M1,Phe, converts the kinetic properties of the M2 into M1-like ones in terms of the response to Mg^<2+>. Thus, it is most likely that the amino acid residue at position 389 plays a key role to confer the Mg^<2+>-conducted regulation. A further investigation would be required to explore the biological significance of such a type of the regulating mechanism.
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