title: The impact of ions on allosteric functions in human liver pyruvate kinase.
abstract: Experimental designs used to monitor the magnitude of an allosteric response can greatly influence observed values. We report here the impact of buffer, monovalent cation, divalent cation, and anion on the magnitude of the allosteric regulation of the affinity of human liver pyruvate kinase (hL-PYK) for substrate, phosphoenolpyruvate (PEP). The magnitudes of the allosteric activation by fructose-1,6-bisphosphate (Fru-1,6-BP) and the allosteric inhibition by alanine are independent of most, but not all buffers tested. However, these magnitudes are dependent on whether Mg(2+) or Mn(2+) is included as the divalent cation. In the presence of Mn(2+), any change in K(app-PEP) caused by Fru-1,6-BP is minimal. hL-PYK activity does not appear to require monovalent cation. Monovalent cation binding in the active site impacts PEP affinity with minimum influence on the magnitude of allosteric coupling. However, Na(+) and Li(+) reduce the magnitude of the allosteric response to Fru-1,6-BP, likely due to mechanisms outside of the active site. Which anion is used to maintain a constant monovalent cation concentration also influences the magnitude of the allosteric response. The value of determining the impact of ions on allosteric function can be appreciated by considering that representative structures used in comparative studies have often been determined using protein crystals grown in diverse buffer and salt conditions.
authors: Department of Biochemistry and Molecular Biology, The University of Kansas Medical Center, Kansas City, Kansas, USA.
authors: Aron W
authors: AW
authors: Fenton
authors: Aileen Y
authors: AY
authors: Alontaga
references: J Biol Chem. 2005 Nov 11;280(45):37924-9
references: 16147999
references: Biochemistry. 1972 Feb 29;11(5):864-78
references: 5059892
references: Methods Enzymol. 2004;380:187-203
references: 15051338
references: Biochemistry. 1977 Apr 5;16(7):1343-50
references: 557338
references: Biochemistry. 1994 Mar 8;33(9):2635-43
references: 8117726
references: Mol Cell Biochem. 1981 Mar 13;35(2):65-75
references: 7015112
references: Biochemistry. 2001 Oct 30;40(43):13097-106
references: 11669648
references: Biochemistry. 2009 May 12;48(18):3816-8
references: 19320443
references: Biophys Chem. 1988 Jun;30(2):159-72
references: 3416042
references: J Biol Chem. 1973 Sep 25;248(18):6436-42
references: 4354210
references: Curr Opin Struct Biol. 2003 Dec;13(6):748-57
references: 14675554
references: Arch Biochem Biophys. 1997 Dec 15;348(2):262-7
references: 9434737
references: Biochemistry. 1998 May 12;37(19):6967-74
references: 9578583
references: Nat Struct Mol Biol. 2006 Sep;13(9):831-8
references: 16906160
references: Science. 2001 Mar 23;291(5512):2429-33
references: 11264542
references: Proteins. 2004 Nov 15;57(3):433-43
references: 15382234
references: Physiol Rev. 2006 Oct;86(4):1049-92
references: 17015484
references: Ital J Biochem. 1979 Sep-Oct;28(5):345-61
references: 399956
references: Arch Biochem Biophys. 1984 Jul;232(1):202-13
references: 6742850
references: Fed Proc. 1982 Nov;41(13):2961-5
references: 7140996
references: IUBMB Life. 2006 Jan;58(1):31-8
references: 16540430
references: Arch Biochem Biophys. 2009 Apr 1;484(1):16-23
references: 19467627
references: Comp Biochem Physiol B Biochem Mol Biol. 2003 Jun;135(2):197-218
references: 12798932
references: J Biol Chem. 1987 Oct 15;262(29):14366-71
references: 3654663
references: Biochemistry. 2003 Jun 3;42(21):6453-9
references: 12767227
references: Int J Biochem. 1978;9(11):785-93
references: 367845
references: J Biol Chem. 1953 Feb;200(2):669-82
references: 13034826
references: Arch Biochem Biophys. 1958 Dec;78(2):477-93
references: 13618030
references: Protein Sci. 2008 Aug;17(8):1295-307
references: 18560010
references: Arch Biochem Biophys. 1997 Sep 15;345(2):199-206
references: 9308890
references: Arch Biochem Biophys. 1971 Mar;143(1):232-9
references: 5561746
references: Annu Rev Biophys Biophys Chem. 1988;17:451-79
references: 3293595
references: Curr Opin Struct Biol. 2006 Feb;16(1):102-8
references: 16423525
references: Arch Biochem Biophys. 1983 Jul 1;224(1):389-401
references: 6870263
references: Biochemistry. 2001 Oct 30;40(43):13088-96
references: 11669647
references: Proteins. 2006 Jun 1;63(4):846-67
references: 16475196
references: Trends Biochem Sci. 2008 Sep;33(9):420-5
references: 18706817
references: Science. 1999 Oct 8;286(5438):295-9
references: 10514373
references: Biochemistry. 2006 May 2;45(17):5421-9
references: 16634623
references: Biochemistry. 1966 Jan;5(1):131-9
references: 5938930
references: Nat Struct Biol. 2003 Jan;10(1):59-69
references: 12483203
references: Biochemistry. 1997 Jun 3;36(22):6803-13
references: 9184163
references: Biochemistry. 1997 Jun 3;36(22):6792-802
references: 9184162