大学精品课件：专业外语The Structure and Function of Protein.docx

1、The Structure and Function of Protein Proteins are the most abundant macromolecules in living cells and constitute 50 percent or more of their dry weight. They are found in all cells and all parts of cells. Proteins also occur in great variety; hundreds of different kinds may be found in a single ce

2、ll. Moreover, proteins have many different biological roles since they are the molecular instruments through with genetic information is expressed. It is therefore appropriate to begin the study of biological macromolecules with the proteins, whose name means “first” or “foremost.” The key to the st

3、ructure of the thousands of different proteins is the group of relatively simple building-block molecules from which proteins are built. All proteins whether from the most ancient lines of bacteria or from the highest forms of life, are constructed from the same basic set of 20 amino acids, covalent

4、ly linked in characteristic sequences. Because each of these amino acids has a distinctive side chain which lends it chemical individuality, this group of 20 building-block molecules may be regarded as the alphabet of protein structure. In this paper we shall also examine peptides, short chains of t

5、wo or more amino acids joined by covalent bonds. What is most remarkable is that cells can join the 20 amino acids in many different combinations and sequences, yielding peptides and proteins having strikingly different properties and activities. From these building blocks different organisms can ma

6、ke such widely diverse products as enzymes, hormones, the lens protein of the eye, feathers, spider webs, tortoise shell, nutritive milk proteins, enkephalins (the bodys own opiates), antibiotics, mushroom poisons, and many other substances having specific biological activity. Amino Acids Have Commo

7、n Structural Features When proteins are boiled with strong acid or base, their amino acid building blocks are released from the covalent linkages that join them into chains. The free amino acids so formed are relatively small molecules, and their structures are all known. The first amino acid to be

8、discovered was asparagines, in 1806. The last of the 20 to be found, threonine, was not identified until 1938. All the amino acids have trivial or common names, sometimes derived from the source from which they were first isolated. Asparagines was first found in asparagus, as one might guess; glutam

9、ic acid was found in wheat gluten; and glycine (Greek, glykos, “sweet”) was so named because of its sweet taste. All of the 20 amino acids found in proteins have as common denominators a carboxyl group and an amino group bonded to the same carbon atom. They differ from each other in their side chain

10、s, or R groups, which vary in structure, size, electric charge, and solubility in water. The 20 amino acids of proteins are often referred to as the standard, primary, or normal amino acids, to distinguish them from other kinds of amino acids present in living organisms but not in proteins. The stan

11、dard amino acids have been assigned three-letter abbreviations and one-letter symbols, which are used as shorthand to indicate to composition and sequence of amino acids in polypeptide chains. Nearly All Amino Acids Have an Asymmetric Carbon Atom We note that all the standard amino acids except one

12、have an asymmetric carbon atom, the carbon, to which are bonded four different substituent groups, i.e., a carboxyl group, an amino group, an R group, and a hydrogen atom. The asymmetric carbon atom is thus a chiral center. As we have seen, compounds with a chiral center occur in two different isome

13、ric forms, which are identical in all chemical and physical properties except one, the direction in which they can cause the rotation of plane-polarized light in a polarimeter. With the single exception of glycine, which has no asymmetric carbon atom, all of the 20 amino acids obtained from the hydr

14、olysis of proteins under sufficiently mild conditions are optically active; i.e., they can rotate the plane-polarized light in one direction or the other. Because of the tetrahedral arrangement of the valence bonds around the carbon atom of amino acids the four different substituent groups can occup

15、y two different arrangements in space, which are nonsuperimposable, mirror images of each other. These two forms are called optical isomers, enantiomers, or stereoisomers. A solution of one stereoisomer of a given amino acid will rotate plane-polarized light to the left (counterclockwise) and is cal

16、led the levorotatory isomer designated (-); the other stereoisomer will rotate plane-polarized light to the same extent but to the right (clockwise) and is called the dextrorotatory isomer designated (+). An equimolar mixture of the (+) and (-) forms will not rotate plane-polarized light. Because al

17、l the amino acids (except glycine) when carefully isolated from proteins do rotate planepolarized light, they evidently occur in only one of their stereoisomeric forms in protein molecules. Optical activity of a stereoisomer is expressed quantitatively by its specific rotation, determined from measu

18、rements of the degree of rotation of a solution of the pure stereoisomer at a given concentration in a tube of a given length in a polarimeter: mLgionconcentratdmoftubelength rotationobserved C D /, deg, 25 the abbreviation dm stands for decimeters (0.1m). The temperature and the wavelength of the l

19、ight employed (usually the D line of sodium, 598nm) must be specified. For the specific rotation of several amino acids, some are levorotatory and others dextrorotatory. Periodic Structures: The Alpha Helix, Beta Pleated Sheet, and Collagen Helix Can a polypeptide chain fold into a regularly repeati

20、ng structure? To answer this question, Pauling and Corey evaluated a variety of potential polypeptide conformations by building precise molecular models of them. They adhered closely to the experimentally observed bond angles and distances for amino acids and small peptides. In 1951, they proposed t

21、wo periodic polypeptide structures, called helix and pleated sheet. The helix is a rod-like structure. The tightly coiled polypeptide main chain forms the inner part of the rod, and the side chains extend outward in a helical array. The helix is stabilized by hydrogen bonds between the NH and CO gro

22、ups of the main chain. The CO group of each amino acid is hydrogen bonded to the NH group of the amino acid that is situated four residues ahead in the linear sequence. Thus, all the main-chain CO and NH groups are hydrogen bonded. Each residue is related to the next one by a translation of 1.5 alon

23、g the helix axis and a rotation of 100 ，which gives 3.6 amino acid residues per turn of helix. Thus, amino acids spaced three and four apart in the linear sequence are spatially quite close to one another in an helix. In contrast, amino acids two apart in the linear sequence are situated on opposite

24、 sides of the helix and so are unlikely to make contact. The pitch of the helix is 5.4 , the product of the translation (1.5 ) and the number of residues per turn (3.6). The screw-sense of a helix can be right-handed (clockwise) or left-handed (counterclockwise); the helices found in proteins are ri

25、ght-handed. The -helix content of proteins of known three-dimensional structure is highly variable. In some, such as myoglobin and hemoglobin, the helix is the major structural motif. Other proteins, such as the digestive enzyme chymotrypsin, are virtually devoid of helix. The single-stranded helix

26、discussed above is usually a rather short rod, typically less than 40 in length. A variation of the -helical theme is used to construct much longer rods, extending to 1000 or more. Two or more helices can entwine around each other to from a cable. Such -helical coiled coils are found in several prot

27、eins: keratin in hair, myosin and tropomyosin in muscle, epidermin in skin, and fibrin in blood clots. The helical cables in these proteins serve a mechanical role in forming stiff bundles of fibers. The structure of the helix was deduced by Pauling and Corey six years before it was actually to be s

28、een in the X-ray reconstruction of the structure of myoglobin. The elucidation of the structure of the helix is a landmark in molecular biology because it demonstrated that the conformation of a polypeptide chain can be predicted if the properties of its components are rigorously and precisely known

29、. In the same year, Pauling and Corey discovered another periodic structural motif, which they named the pleated sheet ( because it was the second structure they elucidated, the helix having been the first). The pleated sheet differs markedly from the helix in that it is a sheet rather than a rod. T

30、he polypeptide chain in the pleated sheet is almost fully extended rather than being tightly coiled as in the helix. The axial distance between adjacent amino acids is 3.5 in contrast with 1.5 for the helix. Another difference is that the pleated sheet is stabilized by hydrogen bonds between NH and

31、CO groups in different polypeptide strands, whereas in the helix the hydrogen bonds are between NH and CO groups in the same polypeptide chain. Adjacent strands in a pleated sheet can run in the same direction (parallel sheet) or in opposite directions (antiparallel sheet). For example, silk fibroin

32、 consists almost entirely of stacks of antiparallel sheets. Such -sheet regions are a recurring structural motif in many proteins. Structural units comprising from two to five parallel or antiparallel strands are especially common. The collagen helix, a third periodic structure, will be discussed in

33、 detail. This specialized structure is responsible for the high tensile strength of collagen, the major component of skin, bone, and tendon. Polypeptide Chains Can Reverse Direction by Making -Turns Most proteins have compact, globular shapes due to frequent reversals of the direction of their polyp

34、eptide chains. Analyses of the three-dimensional structures of numerous proteins have revealed that many of these chain reversals are accomplished by a common structural element called the -turn. The essence of this hairpin turn is that the CO group of residue n of a polypeptide is hydrogen bonded t

35、o the NH group of reside (n+3). Thus, a polypeptide chain can abruptly reverse its direction. Levels of Structure in Protein Architecture In discussing the architecture of proteins, it is convenient to refer to four levels of structure. Primary structure is simply the sequence of amino acids and loc

36、ation of disulfide bridges, if there are any. The primary structure is thus a complete description of the covalent connections of a protein. Secondary structure refers to the steric relationship of amino acid residues that are close to one another in the linear sequence. Some of these steric relatio

37、nships are of a regular kind, giving rise to a periodic structure. The helix, the pleated sheet, and the collagen helix are examples of secondary structure. Tertiary structure refers to the steric relationship of amino acid residues that are far apart in the linear sequence. It should be noted that

38、the dividing line between secondary and tertiary structure is arbitrary. Proteins that contain more than one polypeptide chain display an additional level of structural organization, namely quaternary structure, which refers to the way in which the chains are packed together. Each polypeptide chain

39、in such a protein is called a subunit. Another useful term is domain, which refers to a compact, globular unit of protein structure. Many proteins fold into domains having masses that range from 10 to 20 kdal. The domains of large proteins are usually connected by relatively flexible regions of poly

40、peptide chain. Amino Acid Sequence Specifies Three-Dimensional Structure Insight into the relation between the amino acid sequence of a protein and its conformation came from the work of Christian Anfinsen on ribonuclease, an enzyme that hydrolyzes RNA. Ribonuclease is a single polypeptide chain con

41、sisting of 124 amino acid residues. It contains four disulfide bonds, which can be irreversibly oxidized by performic acid to give cysteic acid residues. Alternatively, these disulfide bonds can be cleaved reversibly by reducing them with a reagent such as -mercaptoethanol, which forms mixed disulfi

42、des with cysteine side chains. In the presence of a large excess of -mercaptoethanol, the mixed disulfides also are reduced, so that the final product is a protein in which the disulfides (cystines) are fully converted into sulfhydryls (cysteines). However, it was found that ribonuclease at 37 and p

43、H7 cannot be readily reduced by -mercaptoethanol unless the protein is partially unfolded by denaturing agents such as urea or guanidine hydrochloride. Although the mechanism of action of these denaturing agents is not fully understood, it is evident that they disrupt noncovalent interactions. Polyp

44、eptide chains devoid of cross-links usually assume a random-coil conformation in 8M urea or 6M guanidine HC1, as evidenced by physical properties such as viscosity and optical rotary spectra. When ribonuclease was treated with -mercaptoethanol in 8M urea, the product was a fully reduced, randomly co

45、iled polyepeptide chain devoid of enzymatic activity. In other words, ribonuclease was denatured by this treatment. Anfinsen then made the critical observation that the denatured ribonulease, freed of urea and -mercaptoethanol by dialysis, slowly regained enzymatic activity. He immediately perceived

46、 the significance of this chance finding: the sulfhydryls of the denatured enzyme became oxidized by air and the enzyme spontaneously refolded into a catalytically active form. Detailed studies then showed that nearly all of the original enzymatic activity was regained if the sulfhydryls were oxidiz

47、ed under suitable conditions. All of the measured physical and chemical properties of the refolded enzyme were virtually identical with those of the native enzyme. These experiments showed that the information needed to specify the complex three-dimensional structure of ribonuclease is contained in

48、its amino acid sequence. Subsequent studies of other proteins have established the generality of this principle, which is a central one in molecular biology: sequence specifies conformation. A quite different result was obtained when reduced ribonuclease was reoxidized while it was still in 8M urea,

49、 This preparation was then dialyzed to remove the urea. Ribonuclease reoxidized in this way had only 1% of the enzymatic activity of the native protein. Why was the outcome of the experiment different from the one in which reduced ribonuclease was reoxidized in a solution free of urea? The reason is that wrong disulfide pairings were formed when the random-coil form of the reduced molecule was reoxidized. There are 105 different ways of pairing eight cysteines to form four disulfides; only o