P-Barrels

The goal of any statistical analysis is inference concerning whether on the basis of available data, some hypothesis about the natural world is true. The hypothesis may consist of the value of some parameter or parameters, such as a physical constant or the exact proportion of an allelic variant in a human population, or the hypothesis may be a qualitative statement, such as This protein adopts an a/p barrel fold or I am currently in Philadelphia. The parameters or hypothesis can be unobservable or as yet unobserved. How the data arise from the parameters is called the model for the system under study and may include estimates of experimental error as well as our best understanding of the physical process of the system. 

The eight-stranded a/p-barrel stmcture is one of the largest and most regular of all domain stmctures. A minimum of about 200 residues are required to form this structure. It has been found in many different proteins, most of which are enzymes, with completely different amino acid sequences and... 

Branched hydrophobic side chains dominate the core of oi/p barrels... 

The packing interactions between a helices and p strands are dominated by the residues Val (V), He (I), and Leu (L), which have branched hydrophobic side chains. This is reflected in the amino acid composition these three amino acids comprise approximately 40% of the residues of the P strands in parallel P sheets. The important role that these residues play in packing a helices against P sheets is particularly obvious in a/P-barrel structures, as shown in Table 4.1. 

Figure 4.3 In most a/p-barrel structures the eight p strands of the barrel enclose a tightly packed hydrophobic core formed entirely by side chains from the p strands. The core is arranged in three layers, with each layer containing four side chains from alternate p strands. The schematic diagram shows this packing arrangement in the a/p barrel of the enzyme glycolate oxidase, the structure of which was determined by Carl Branden and colleagues in Uppsala, Sweden.

There is one exception to the rule that requires bulky hydrophobic residues to fill the interior of eight-stranded a/p barrels in order to form a tightly packed hydrophobic core. The coenzyme Biz-dependent enzyme methylmalonyl-coenzyme A mutase, the x-ray structure of which was determined by Phil Evans and colleagues at the MRC Laboratory of Molecular... 

Figure 4.4 Schematic diagram of the structure of the a/p-barrel domain of the enzyme methylmalonyl-coenzyme A mutase. Alpha helices are red, and p strands are blue. The inside of the barrel is lined by small hydrophilic side chains (serine and threonine) from the p strands, which creates a hole in the middle where one of the substrate molecules, coenzyme A (green), binds along the axis of the barrel from one end to the other. (Adapted from a computer-generated diagram provided by P. Evans.)...

Figure 4.5 The polypeptide chain of the enzyme pyruvate kinase folds into several domains, one of which is an a/p barrel (red). One of the loop regions in this barrel domain is extended and comprises about 100 amino acid residues that fold into a separate domain (blue) built up from antiparallel P strands. The C-terminal region of about 140 residues forms a third domain (green), which is an open twisted a/p structure.

All known eight-stranded a/p-barrel domains have enzymatic functions that include isomerization of small sugar molecules, oxidation by flavin coenzymes, phosphate transfer, and degradation of sugar polymers. In some of these enzymes the barrel domain comprises the whole subunit of the protein in others the polypeptide chain is longer and forms several additional domains. An enzymatic function in these multidomain subunits, however, is always associated with the barrel domain. 

PRA-isomerase lGP-synthase, a bifunctional enzyme from E. coli that catalyzes two reactions in the synthesis of tryptophan (Figure 4.6), has a polypeptide chain that forms two a/p barrels. The stmcture of this enzyme, solved at 2.8 A in the laboratory of Hans Jansonius in Basel, Switzerland, showed that residues 48-254 form one barrel with IGP-synthase activity, while residues 255-450 form the second barrel with PRA-isomerase activity (Figure 4.7). 

Figure 4.7 Two of the enzymatic activities involved in the biosynthesis of tryptophan in E. coli, phosphoribosyl anthranilate (PRA) isomerase and indoleglycerol phosphate (IGP) synthase, are performed by two separate domains in the polypeptide chain of a bifunctional enzyme. Both these domains are a/p-barrel structures, oriented such that their active sites are on opposite sides of the molecule. The two catalytic reactions are therefore independent of each other. The diagram shows the IGP-synthase domain (residues 48-254) with dark colors and the PRA-isomerase domain with light colors. The a helices are sequentially labeled a-h in both barrel domains. Residue 255 (arrow) is the first residue of the second domain. (Adapted from J.P. Priestle et al., Proc.

Figure 4.8 The active site in all a/p barrels is in a pocket formed by the loop regions that connect the carboxy ends of the p strands with the adjacent a helices, as shown schematically in (a), where only two such loops are shown, (b) A view from the top of the barrel of the active site of the enzyme RuBisCo (ribulose bisphosphate carboxylase), which is involved in CO2 fixation in plants. A substrate analog (red) binds across the barrel with the two phosphate groups, PI and P2, on opposite sides of the pocket. A number of charged side chains (blue) from different loops as welt as a Mg ion (yellow) form the substrate-binding site and provide catalytic groups. The structure of this 500 kD enzyme was determined to 2.4 A resolution in the laboratory of Carl Branden, in Uppsala, Sweden. (Adapted from an original drawing provided by Bo Furugren.)...

We have described a general relationship between structure and function for the a/p-barrel structures. They all have the active site at the same position with respect to their common structure in spite of having different functions as well as different amino acid sequences. We can now ask if similar relationships also occur for the open a/p-sheet structures in spite of their much greater variation in structure. Can the position of the active sites be predicted from the structures of many open-sheet a/p proteins ... 

In almost every one of the more than 100 different known a/p structures 1 of this class the active site is at the carboxy edge of the p sheet. Functional residues are provided by the loop regions that connect the carboxy end of the strands with the amino end of the a helices. In this one respect a fun-I damental similarity therefore exists between the a/p-barrel structures and the I open a/p-sheet structures. 

The a/p-barrel structure is one of the largest and most regular of all domain structures, comprising about 250 amino acids. It has so far been found in more than 20 different proteins, with completely different amino acid sequences and different functions. They are all enzymes that are modeled on this common scaffold of eight parallel p strands surrounded by eight a helices. They all have their active sites in very similar positions, at the bottom of a funnel-shaped pocket created by the loops that connect the carboxy end of the p strands with the amino end of the a helices. The specific enzymatic activity is, in each case, determined by the lengths and amino acid sequences of these loop regions which do not contribute to the stability of the fold. 

Easters, I., et al. Structural principles of parallel p barrels in proteins. Proc. Natl. Acad. Sci. USA 85 3338-3342, 1988. 

Easters, I., Wodak, S.J., Pio, F. The design of idealized a/p-barrels analysis of p-sheet closure requirements. Proteins 7 249-256, 1990. 

Lesk, A.M., Branden, C.-L, Chothia, C. Structural principles of a/p barrel proteins the packing of the interior of the sheet. Proteins 5 139-148, 1989. 

The simplest topology is obtained if each successive p strand is added adjacent to the previous strand until the last strand is joined by hydrogen bonds to the first strand and the barrel is closed (Figure 5.2). These are called up-and-down P sheets or barrels. The arrangement of p strands is similar to that in the a/P-barrel structures we have just described in Chapter 4, except that here the strands are antiparallel and all the connections are hairpins. The structural and functional versatility of even this simple arrangement will be illustrated by two examples. 

Figure S.3 Schematic diagram of the structure of human plasma retinol-binding protein (RBP), which is an up-and-down P barrel. The eight antiparallel P strands twist and curl such that the structure can also be regarded as two p sheets (green and blue) packed against each other. Some of the twisted p strands (red) participate in both P sheets. A retinol molecule, vitamin A (yellow), is bound inside the barrel, between the two P sheets, such that its only hydrophilic part (an OH tail) is at the surface of the molecule. The topological diagram of this stmcture is the same as that in Figure 5.2. (Courtesy of Alwyn Jones, Uppsala, Sweden.)...

There is a second family of small lipid-binding proteins, the P2 family, which include among others cellular retinol- and fatty acid-binding proteins as well as a protein, P2, from myelin in the peripheral nervous system. However, members of this second family have ten antiparallel p strands in their barrels compared with the eight strands found in the barrels of the RBP superfamily. Members of the P2 family show no amino acid sequence homology to members of the RBP superfamily. Nevertheless, their three-dimensional structures have similar architecture and topology, being up-and-down P barrels. 

We saw in Chapter 2 that the Greek key motif provides a simple way to connect antiparallel p strands that are on opposite sides of a barrel structure. We will now look at how this motif is incorporated into some of the simple antiparallel P-barrel structures and show that an antiparallel P sheet of eight strands can be built up only by hairpin and/or Greek key motifs, if the connections do not cross between the two ends of the p sheet. 

Figure 9.18 Schematic digram of the structure of the DNA-binding domain of p53. (a) The DNA binding domain of p53 folds into an antiparallel p barrel with long loop regions—...

The polypeptide chain is folded into two domains (Figure 11.7), each of which contains about 120 amino acids. The two domains are both of the antiparallel p-barrel type, each containing six p strands with the same topology (Figure 11.8). Even though the actual structure looks complicated, the topology is very simple, a Greek key motif (strands 1-4) followed by an antiparallel hairpin motif (strands 5 and 6). 

Figure 11.8 Topology diagrams of the domain structure of chymotrypsin. The chain is folded into a six-stranded antiparallel p barrel arranged as a Greek key motif followed by a hairpin motif.

---