H. Lodish - Molecular Cell Biology (5ed, Freeman, 2003) (796244), страница 27
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This view represents a proteinas it is “seen” by another molecule.Motifs Are Regular Combinations of SecondaryStructuresParticular combinations of secondary structures, called motifs or folds, build up the tertiary structure of a protein. Insome cases, motifs are signatures for a specific function. Forexample, the helix-loop-helix is a Ca2-binding motifmarked by the presence of certain hydrophilic residues at invariant positions in the loop (Figure 3-6a). Oxygen atoms in63representations of the structure of Ras,a monomeric guanine nucleotide-bindingprotein. The inactive, guanosinediphosphate (GDP)–bound form is shownin all four panels, with GDP always depictedin blue spacefill.
(a) The C backbone tracedemonstrates how the polypeptide ispacked into the smallest possible volume.(b) A ball-and-stick representation revealsthe location of all atoms. (c) A ribbonrepresentation emphasizes how strands(blue) and helices (red) are organized inthe protein. Note the turns and loopsconnecting pairs of helices and strands.(d) A model of the water-accessible surfacereveals the numerous lumps, bumps, andcrevices on the protein surface. Regions ofpositive charge are shaded blue; regions ofnegative charge are shaded red.the invariant residues bind a Ca2 ion through ionic bonds.This motif, also called the EF hand, has been found in morethan 100 calcium-binding proteins.
In another commonmotif, the zinc finger, three secondary structures—an helixand two strands with an antiparallel orientation—form afingerlike bundle held together by a zinc ion (Figure 3-6b).This motif is most commonly found in proteins that bindRNA or DNA.Many proteins, especially fibrous proteins, self-associateinto oligomers by using a third motif, the coiled coil. In theseproteins, each polypeptide chain contains -helical segmentsin which the hydrophobic residues, although apparentlyrandomly arranged, are in a regular pattern—a repeatedheptad sequence.
In the heptad, a hydrophobic residue—sometimes valine, alanine, or methionine—is at position 1and a leucine residue is at position 4. Because hydrophilicside chains extend from one side of the helix and hydrophobic side chains extend from the opposite side, the overall helical structure is amphipathic. The amphipathic character ofthese helices permits two, three, or four helices to windaround each other, forming a coiled coil; hence the name ofthis motif (Figure 3-6c).We will encounter numerous additional motifs in laterdiscussions of other proteins in this chapter and other chapters. The presence of the same motif in different proteinswith similar functions clearly indicates that these useful64CHAPTER 3 • Protein Structure and Function(c) Coiled coil motifN(a) Helix-loop-helix motifCa2+N(b) Zinc-finger motifLeu (4)AsnAspCThrHisZn2+Val (1)H2OGluAspCysNLeu (4)HisAsn (1)CysLeu (4)Val (1)NLeu (4)CConsensus sequence:F/Y - C - - C - - - - F/Y - - - - - - - - H - - - H CConsensus sequence:D/N - D/N - D/N/S - [backbone O] - - - - E/D▲ FIGURE 3-6 Motifs of protein secondary structure.(a) Two helices connected by a short loop in a specificconformation constitute a helix-loop-helix motif.
This motif existsin many calcium-binding and DNA-binding regulatory proteins.In calcium-binding proteins such as calmodulin, oxygen atomsfrom five loop residues and one water molecule form ionic bondswith a Ca2 ion. (b) The zinc-finger motif is present in manyDNA-binding proteins that help regulate transcription. A Zn2 ionis held between a pair of strands (blue) and a single helix(red) by a pair of cysteine residues and a pair of histidineresidues.
The two invariant cysteine residues are usually atpositions 3 and 6 and the two invariant histidine residues arecombinations of secondary structures have been conserved inevolution. To date, hundreds of motifs have been catalogedand proteins are now classified according to their motifs.Structural and Functional Domains Are Modulesof Tertiary StructureThe tertiary structure of proteins larger than 15,000 MW istypically subdivided into distinct regions called domains.Structurally, a domain is a compactly folded region ofpolypeptide. For large proteins, domains can be recognizedin structures determined by x-ray crystallography or in images captured by electron microscopy.
Although these discrete regions are well distinguished or physically separatedfrom one another, they are connected by intervening segments of the polypeptide chain. Each of the subunits inhemagglutinin, for example, contains a globular domain anda fibrous domain (Figure 3-7a).CHeptad repeat:[V/N/M] - - L - - -at positions 20 and 24 in this 25-residue motif.
(c) The paralleltwo-stranded coiled-coil motif found in the transcription factorGcn4 is characterized by two helices wound around oneanother. Helix packing is stabilized by interactions betweenhydrophobic side chains (red and blue) present at regularintervals along the surfaces of the intertwined helices. Each helix exhibits a characteristic heptad repeat sequence with ahydrophobic residue at positions 1 and 4. [See A. Lewit-Bentleyand S. Rety, 2000, EF-hand calcium-binding proteins, Curr.
Opin. Struct.Biol. 10:637–643; S. A. Wolfe, L. Nekludova, and C. O. Pabo, 2000,DNA recognition by Cys2His2 zinc finger proteins, Ann. Rev. Biophys.Biomol. Struct. 29:183–212.]A structural domain consists of 100–150 residues in various combinations of motifs. Often a domain is characterizedby some interesting structural feature: an unusual abundanceof a particular amino acid (e.g., a proline-rich domain, anacidic domain), sequences common to (conserved in) manyproteins (e.g., SH3, or Src homology region 3), or a particular secondary-structure motif (e.g., zinc-finger motif in thekringle domain).Domains are sometimes defined in functional terms onthe basis of observations that an activity of a protein is localized to a small region along its length. For instance, a particular region or regions of a protein may be responsible forits catalytic activity (e.g., a kinase domain) or binding ability(e.g., a DNA-binding domain, a membrane-binding domain).Functional domains are often identified experimentally bywhittling down a protein to its smallest active fragment withthe aid of proteases, enzymes that cleave the polypeptidebackbone.
Alternatively, the DNA encoding a protein can be3.1 • Hierarchical Structure of Proteins(a)(b)Sialic acidPROXIMALGlobulardomainFibrousdomainNHA1NCViralmembranesubjected to mutagenesis so that segments of the protein’sbackbone are removed or changed. The activity of the truncated or altered protein product synthesized from the mutated gene is then monitored and serves as a source of insightabout which part of a protein is critical to its function.The organization of large proteins into multiple domains illustrates the principle that complex molecules arebuilt from simpler components. Like motifs of secondarystructure, domains of tertiary structure are incorporated asmodules into different proteins.
In Chapter 10 we considerthe mechanism by which the gene segments that correspondto domains became shuffled in the course of evolution, resulting in their appearance in many proteins. The modularapproach to protein architecture is particularly easy to recognize in large proteins, which tend to be mosaics of different domains and thus can perform different functionssimultaneously.The epidermal growth factor (EGF) domain is one example of a module that is present in several proteins (Figure 3-8).EGF is a small, soluble peptide hormone that binds to cells inthe embryo and in skin and connective tissue in adults, causing them to divide.
It is generated by proteolytic cleavage between repeated EGF domains in the EGF precursor protein,which is anchored in the cell membrane by a membranespanning domain. EGF modules are also present in otherproteins and are liberated by proteolysis; these proteins include tissue plasminogen activator (TPA), a protease that isused to dissolve blood clots in heart attack victims; FIGURE 3-7 Tertiary and quaternarylevels of structure in hemagglutinin (HA),a surface protein on influenza virus. Thislong multimeric molecule has three identicalsubunits, each composed of two polypeptidechains, HA1 and HA2.
(a) Tertiary structure ofeach HA subunit constitutes the folding of itshelices and strands into a compact structurethat is 13.5 nm long and divided into twodomains. The membrane-distal domain isfolded into a globular conformation. Themembrane-proximal domain has a fibrous,stemlike conformation owing to the alignmentof two long helices (cylinders) of HA2 with strands in HA1. Short turns and longerloops, which usually lie at the surface of themolecule, connect the helices and strands ina given chain. (b) Quaternary structure of HAis stabilized by lateral interactions betweenthe long helices (cylinders) in the fibrousdomains of the three subunits (yellow, blue,and green), forming a triple-stranded coiledcoil stalk.
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