Class Note for BIOC 462A at UA
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Date Created: 02/06/15
Biochemistry 462a Proteins Tertiary and Quaternary Structure Reading Chapter 6 Practice problems Chapter 6 5 8 Proteins extra problems Tertiary Structure 0 Most proteins are globular essentially spherical 0 They posses both secondary structure and are folded into compact Tertiary Structures 0 From xray diffraction studies we know the details of the three dimensional structure of many globular proteins 0 Every protein has a unique three dimensional structure made up of a variety of helices beta sheets and nonregular regions which are folded in a specific manner 0 Combinations of secondary structural elements are often found in proteins These combinations are called motifs Some motifs have functional signi cance such as the helix loophelix DNAbinding motif or the EF hand calciumbinding motif Others serve only a structural role Domains 0 Polypeptide chains of gt200 amino acids often fold into two or more compact globular clusters called domains 0 There are three main types of domains 0 0L domains are composed of OLhelices o 5 domains contain antiparallel 5 sheets and usually contain two 5 sheets packed against each other 0 UJB domains contain the B OL B motif of parallel 5 sheets 0 Adjacent domains are connected by one or two segments of the polypeptide chain Generalizations about Protein Structure Examination of the structure of many proteins has lead to some important generalizations about the folding of the polypeptide chain in protein tertiary structures 0 All Globular Proteins have a def39med Inside and Outside Examination of the amino acid sequence of proteins shows no obvious pattern of hydrophobic and hydrophilic amino acids but in the tertiary structure of proteins almost all the hydrophobic sidechains are found in the interior of the protein and almost all hydrophilic sidechains are found on the outside of the protein interacting with water 0 Globular Proteins are compact There is no space inside so that water is effectively excluded from the hydrophobic interior 0 Nearly all buried Hydrogen Bond Donors eg Ser form hydrogen bonds with Hydrogen Bond Acceptors eg Gln In essence hydrogen bond formation neutralizes the polarity of the hydrogenbonding group 0 B sheets are usually Twisted or Wrapped into Barrel Structures 0 Loops and Turns are on the outside of the protein o Mutations which place a hydrophobic sidechain on the surface can cause signi cant changes in the folding of the protein In this example Arc protein a B sheet is converted to a helix by changing the amino acid sequence of the B sheet from G1nPheAsn LeuArg Trp to Gln PheLeu AsnArg Trp In the B sheet the interchange of Leu and Asn would cause the Leu to stick into the solvent and the Asn to stick into the interior of the protein However the secondary structure is reorganized to form an unusual righthanded helix which moves the Leu from a surface to an interior position and the Asn from an interior to a surface position Stabilization of the Tertiary Structure 0 Each protein has a unique tertiary structure THE NATIVE STATE which is de ned by a unique arrangement of all the atoms in the molecule 0 There are of course many other possible arrangements of these atoms but these other arrangements do not posses biological activity and collectively they are termed THE DENATURED STATE 0 What factors favor the native state over the denatured state 0 More precisely why is AG for the conversion of the denatured state to the native state favorable AG AHTAS so we need to consider both entropic effect AS and enthalpic effects AH Entropy Effects 0 In the denatured state the hydrophobic sidechains are in contact with water which is an unfavorable situation 0 Because water cannot interact favorably with hydrophobic groups no H bonds are possible the water must reorganize itself in order to accommodate such a group clathrate formation 0 This is an entropically unfavorable process 0 When the denatured state is converted to the native state the hydrophobic sidechains are buried in the interior of the protein away from water 0 This allows the previously ordered water molecules to becomes disordered an entropically favorable process 0 The is known as the HYDROPHOBIC EFFECT and contributes a favorable entropy to the formation of the native state 0 Opposing this is the unfavorable conformational entropy associated with formation of the native state which arises from the fact that the native state represents a single unique conformation of the protein whereas the denatured state represents many different conformations Enthalpy Effects 0 These arise from favorable sidechain group interactions in the native state 0 Dipoledipole van der Waals forces and dipoleinduced dipole forces London dispersion forces are particularly important in the core of the protein where the dielectric constant is low and the atoms of the sidechains are packed closely together Indeed these forces are the major source of the favorable enthalpy associated with protein folding Electrostatic interactions and hydrogen bonding play important roles in stabilizing the native state although they contribute little to the energetics This seemingly paradoxical statement can be understood as follows 0 All potential hydrogen bond donors and acceptors in a protein will form hydrogen bonds whether in the native or denatured state In the denatured state the hydrogen bonds will most often be with water but in the native state there will be specific hydrogen bonds formed the hydrogen bonds that stabilize OLhelices or Bsheets or that are formed between buried polar groups Because there is little difference between the energy of a hydrogen bond between a CO and water or between a CO and HN there is not much energy involved in forming a helix or a sheet However the formation of these SPECIFIC hydrogen bonds is absolutely essential for the correct folding of the polypeptide chain into the native state A similar argument applies to electrostatic interactions salt bridges Formation of salt bridges involves desolvation of the ion unfavorable followed by electrostatic interaction favorable The net energy difference is small but the formation of SPECIFIC salt bridges is critical in stabilizing the native state For most proteins the favorable free energy of formation of the native state is the difference between large enthalpy and entropy effects in many proteins AH predominates but in some cases AS predominates IProtein IAG IAH IAS lLysozyme 62 220 530 letochrome c I44 I52 I27 IMyoglobin 50 0 170 Disulflde Bonds In proteins that contain them disulfide bonds help to stabilize the native state because the correct formation of disulfide bonds can only occur when the the two SH groups are positioned correctly The Importance of Proline 0 There has been much speculation that the conversion of an OLhelix to a Bsheet may be responsible for the aggregation of proteins into the fibrils that characterize prion disease or Alzheimer39s disease Prolines often serve to delineate the amino terminal end of a helix Is it possible that a mutation that substitutes a Pro for another amino acid in a helix could cause the formation of a Bsheet 0 Recently the reverse change has been observed In this case substitution of an Ala for a Pro changes a Bsheet to an OLhelix WZ Yen et al Protein Science 11875l998 Quaternary Structure Many large proteins contain more than one polypeptide chain SUBUNIT The spatial arrangement of these subunits is the quaternary structure of the protein The forces that hold subunits together are the same as those that stabilize the tertiary structure of proteins van der Waals and London Dispersion forces salt bridges and hydrogen bonds The contact region between subunits resembles the interior of a protein The subunits may be identical or nonidentical but there is always a de ned stoichiometery Several proteins function by forming transient complexes with other proteins For example electron transfer from cytochrome c to cytochrome c peroxidase involves the formation of such a complex Because such complexes are transient the interactions are weaker than those that stabilize the quaternary structure of hemoglobin In the cytochrome ccytochrome c peroxidase complex there are specific electrostatic interactions that guide the two proteins together in the correct orientation Protein Folding The amino acid sequence of a protein contains all the information required for the protein to fold into the correct biologically active threedimensional structure One of the important unsolved problems in biochemistry is the quotfolding problemquot by which we mean quothow do proteins foldquot o The process most likely begins with the formation of secondary structural elements which serve as nucleation foci around which the native structure of the protein can fold o It is likely that OLhelices are the most important nucleation foci because they are formed from amino acids which are next to each other in the linear sequence whereas 5 sheets are often formed by the interaction of strands that are far apart in the linear sequence 0 Nuclei with the proper nativelike secondary structure probably interact with each until they form a nativelike domain 0 Ultimately these secondary structural domains will come together to form a structure with extensive secondary structure but disordered tertiary structure the molten globule state 0 Finally small rearrangements of the molten globule generate the native conformation Whlle rt ls aeeepted that the ammo aerd sequence of a protern eontarns all the lnformatlon neeessary for the protern to adopt rts eorreet threerddmenslonal strueture rt ls beeomrng elear that there are several aeeessory proterns that assrstrn the foldlng proeess Among these are the Mnleeular Chapernnes multlrsubumt proterns that uulrze ATP to gulde proterns along the eorreet fol dmg pathway fonlded Pnneins Whlle most mlsfolded proterns are slmply degraded m the eell there are some srtuatrons m whreh the mlsfoldedproteln aeeumulates m the eell andleads to apathologreal condmon What seems to happen ls that the mlsfolded lntermedlate forms rnappropnate aggregates leadlng to the formauon of large polymers see slckle eell hemoglobln for detarls of one example Among the known examples of dlseases eaused by mlsfolded proterns are eysue brosls seuryy serapre and Alzhelmer s dlsease Prutein Dynamics Pruteins are nnt racks They undergo many complex mouons that are undoubtedly essenual for therr brologreal aeuyrty Another way to get a feellng for the dynamll propemes of a protern ls to analyze the protern strueture uslng N39MR NMR data do not yreld a umque strueture but a range of struetures that are eonsrstentwrth the data The range of those struetures shown here for the baekbone eonformatrons of bovlne panereaue trypsrn rnhrbrtor K D Bemdt P Guntert LP M orbond K Wuthneh JMal8m1 227 757 1992 gyes an lmpresslon of the dynaml propemes othe protern m soluuon
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