Three Dimensional Structure of Proteins Tertiary Structure and Protein Folding Sections 4 3 and 4 4 Biochem 4511 Figures Essentials of Biochemistry 3rd Ed OSU Custom Edition Principles of Biochemistry 5th Ed Moran et al Lehninger Principles of Biochemistry 5th Ed Nelson Cox Fundamentals of Biochemistry 2nd Ed Voet Voet Pratt Common Depictions of Protein Structure Proteins fold into defined structures with differing secondary structure and differing organization helical protein Growth hormone protein flavodoxin protein gb crystallin Protein with very little 2 structure tachystatin Separate structural clusters within one protein chain are Domains Glyceraldehyde 3 phosphate dehydrogenase Src Kinase Assemblies from separate protein chains quaternary structure Nitrite reductase homotrimer Hemoglobin Heterotetramer 2a 2b subunits E Coli fumarase homotetramer Bacterial methane hydroxylase Hexamer Dimer of heterotrimers Subunit Separate protein chain multisubunit has multiple chains Quaternary Structure Protein Complexes Monomer Homoodimer Trimer Heterodimer Tetramer 4 subunits Pentamer 5 subunits Hexamer 6 subunits Octamer 8 subunits Quaternary Structure Hemoglobin Heterotetramer 2a 2b subunits Cooperative subunit interactions and dynamics contributes to regulation of complex biological activity Chapter 5 Virus capsids massive quaternary assemblies Polio virus Tobacco mosaic virus Protein Structure and Stability Surface view of myoglobin blue charged AA yellow hydrophobic AA Cross section of myoglobin Water as a Solvent Water as a solvent provides most of the driving force behind protein folding As a reminder Hydrophilic Water loving Forms favorable interactions with water molecules dissolves well in water Hydrophobic Water fearing hating Does not make favorable interactions with water doesn t dissolve well Amphipathic Molecules and Water Individual lipids Lipid clusters Water is ordered around each molecule Water is excluded and is ordered around border lipids Proteins are Amphipathic Protein backbones are polar Polarity of side chains vary from very hydrophilic to very hydrophobic Proteins fold to maximize intramolecular hydrophobic interactions in aqueous solvent You should be able to predict approximate hydrophobicity ie clusters not numbers from sidechain structure Hydrophobic Effect Stability Difference between folded and unfolded states Folded Buried hydrophobic groups shielded from water Unfolded Unfavorable water cages entropic cost Energetic Benefit of Hydrophobic Effect Each methylene group CH2 removed from aqueous solvent is estimated to provide 3 kJ mol of free energy There are many hydrocarbon chains in a protein By comparison Each hydrogen bond is 20 kJ mol Protein Folding is Entropy Driven Protein folding is PRIMARILY driven by entropy Protein structure and stability is a fine balance between enthalpic and entropic forces Cross links in proteins Covalent disulfide bonds may stabilize protein structures when not in a reducing environment cytoplasm is reducing Erabutoxin a sea snake toxin Lysozyme extracellular In small proteins disulfides may provide significant stability Larger proteins may retain fold in absence of disulfides Cross links in proteins TFIIIA zinc finger Gal4 zinc finger Metal centers can be functional or structural or both Zn2 often structural does not undergo oxidation Interacts with S N O but particularly His Cys Hydrogen Bonds Hydrogen bonds are key to defining protein structure and Hydrogen bonds are necessary to remain stable BUT BECAUSE Hydrogen bonds are not the major force in protein stability Stability is the difference between folded and unfolded states Folded form Intramolecular hydrogen bonds Unfolded form hydrogen bonds with water Hydrogen bonds provide little net benefit DESOLVATION COST Hydrogen bonds are not the primary driving force for protein folding structure Desolvation Cost Forming hydrogen bonds in the protein interior compensates for broken hydrogen bonds to water desolvation cost Polar groups NOT forming hydrogen bonds in a folded protein are energetically unfavorable because the desolvation cost is not recovered Hydrogen Bonds Specificity vs Stability Hydrophobic effect entropy Protein will fold to globular shape or tertiary structure Major stabilizer Hydrogen bonds Minimize unpaired polar groups buried in the hydrophobic core and thus drive what type of globular shape will be formed Specificity Secondary structure helix and sheet Best way to pack the space in folded protein while compensating for lost interactions with water desolvation cost Specificity Ion Pairs and Salt Bridges Electrostatic interactions Charged side chains make specific salt bridges Ion Pairs and Salt Bridges Salt bridge formation 86 kJ mol recovers approximately the same energy as the cost of taking charges out of water entropy in side chain motion enthalpy in solvent interaction Unpaired charges in the interior of a protein are very unfavorable Desolvation Costs Interior salt bridges contribute little stability and a lot of specificity Exterior salt bridges likely contribute little stability or specificity because of solvent water Asp 60 Dipole Dipole Interactions Dipole dipole interactions act over short distances to stabilize particular protein conformations Van der Waals forces Effective only over short distances but energetic benefit in forming a well defined protein core Not the same as hydrophobic effect Strongest Weakest Hydrophobic Forces Van der Waals contacts are specific interactions between hydrophobic groups that stabilize a well folded protein interior Different than hydrophobic effect of releasing entropy cage by burying hydrophobic residues Hydrophobic Forces in Proteins Like hydrogen bonds van der Waals forces provide an enthalpic benefit to the folded protein core However van der Waals forces are NOT as important as the release of ordered water entropy as the driving force for protein folding Most important take home concept Everything else just selects for the most energetically favorable way to bury the hydrophobic side chains specificity Summary Hydrophobic Effect Desolvation Cost Forces that lead to specificity of tertiary protein structure Cross links Disulfide bonds and metal centers Hydrogen bonds Dipole dipole interactions Van Der Waals interactions Electrostatic interactions salt bridges Proteins are Marginally Stable An average 100 residue protein is stable by approximately 40 kJ mol Such a protein will unfold if you add 40 kJ