Thermodynamic analysis of the heparin interaction with a basic cyclic peptide using isothermal titration calorimetry.
Brain natriuretic peptide (BNP) was examined as part of a continuing study of the interaction of proteins and peptides with the glycosaminoglycan heparin. BNP was tentatively identified as a heparin-binding protein on the basis of its cyclic structure and the high frequency of the basic amino acid residues, lysine and arginine. Thermodynamic analysis using isothermal titration calorimetry confirmed heparin binding to BNP with a micromolar Kd. Surprisingly, despite the high frequency (22%) of basic residues in BNP, only a small portion of the free energy of this interaction resulted from ionic contributions under physiologic conditions. The contribution of polar amino acids, representing 28% of BNP, was next examined in a variety of different buffers. These experiments demonstrated the transfer of five protons from buffer to BNP on heparin binding, suggesting that hydrogen bonding between the polar residues of BNP and heparin is a major factor contributing to the free energy of BNP binding to heparin. Hydrophobic forces apparently play only a small role in binding. Heparin contains few nonpolar functional groups, and a positive change in heat capacity (DeltaCp = 1 kcal/mol) demonstrates the loss of polar residues on BNP-heparin binding.
Hileman
R E
RE
Division of Medicinal and Natural Products Chemistry, Department of Chemical and Biochemical Engineering, University of Iowa, Iowa City 52242, USA.
Jennings
R N
RN
Linhardt
R J
RJ
D000595:Amino Acid Sequence; D002021:Buffers; D002151:Calorimetry; D006493:Heparin / Q000737:chemistry* / Q000378:metabolism; D006801:Humans; D006863:Hydrogen-Ion Concentration; D008969:Molecular Sequence Data; D020097:Natriuretic Peptide, Brain / Q000737:chemistry* / Q000378:metabolism; D010456:Peptides, Cyclic / Q000737:chemistry* / Q000378:metabolism; D011108:Polymers / Q000378:metabolism; D011485:Protein Binding / Q000187:drug effects; D012965:Sodium Chloride / Q000494:pharmacology; D013696:Temperature; D013816:Thermodynamics
D002021:Buffers; D010456:Peptides, Cyclic; D011108:Polymers; D020097:Natriuretic Peptide, Brain; D012965:Sodium Chloride; D006493:Heparin