The monomer of pyruvate kinase, subtype M1, is both a kinase and a cytosolic thyroid hormone binding protein.
Using a T7 expression system, the monomer of rat pituitary pyruvate kinase, subtype M1 (PKM1), was overexpressed in Escherichia coli and purified to homogeneity. The monomeric p58-M1 has intrinsic enzymatic activity with a Vmax of 79 +/- 20 units/mg and Km's for ADP and PEP of 1.43 +/- 0.76 and 0.14 +/- 0.07 mM, respectively. The monomer binds 3,3',5-triiodo-L-thyronine (T3) with Ka = 1.5 x 10(7) M-1. The order of analog specificity is L-T3 greater than L-thyroxine greater than D-T3 greater than 3'-isopropyl-3,5-diiodo-L-thyronine greater than or equal to 3',5',3-triiodo-L-thyronine. In contrast, tetrameric PKM1 lacks T3 binding activity. The kinase activity of p58-M1 is inhibited by T3 and its analogs in a concentration-dependent manner with the order of inhibitory activity similar to that of binding activity. This inhibition, however, is reversed by the addition of fructose 1,6-bisphosphate. p58-M1 is the second PK isoenzyme monomer to be identified as having thyroid hormone binding activity.
Parkison
C
C
Laboratory of Molecular Biology, National Cancer Institute, National Institutes of Health, Bethesda, MD 20892.
Ashizawa
K
K
McPhie
P
P
Lin
K H
KH
Cheng
S Y
SY
D000818:Animals; D002460:Cell Line; D003001:Cloning, Molecular; D003600:Cytosol / Q000378:metabolism; D004926:Escherichia coli / Q000235:genetics; D015870:Gene Expression; D006801:Humans; D007527:Isoenzymes / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D007700:Kinetics; D046911:Macromolecular Substances; D008970:Molecular Weight; D010957:Plasmids; D011770:Pyruvate Kinase / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D011988:Receptors, Thyroid Hormone / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D011994:Recombinant Proteins / Q000302:isolation & purification / Q000378:metabolism; D013604:T-Phages / Q000235:genetics; D013963:Thyroid Hormones / Q000378:metabolism
D007527:Isoenzymes; D046911:Macromolecular Substances; D011988:Receptors, Thyroid Hormone; D011994:Recombinant Proteins; D013963:Thyroid Hormones; D011770:Pyruvate Kinase