Two domains

Interestingly, there are many proteins with two domains that show a very clear hinge-bending motion with an obvious functional significance. Such domains have often been reported in the literature, but were never detected on an automated basis. 

Fig. 5. Rigid-body analysis of citrate synthase, using two X-ray structures (after Hayward and Berendsen, Proteins 30 (1998) 144). The decomposition of the protein into two domains (dark gray and white) and two interconnecting regions (light gray) is shown, together with the hinge axis for the closing/opening motion between them.

The catalytic subunit of cAPK contains two domains connected by a peptide linker. ATP binds in a deep cleft between the two domains. Presently, crystal structures showed cAPK in three different conformations, (1) in a closed conformation in the ternary complex with ATP or other tight-binding ligands and a peptide inhibitor PKI(5-24), (2) in an intermediate conformation in the binary complex with adenosine, and (3) in an open conformation in the binary complex of mammalian cAPK with PKI(5-24). Fig.l shows a superposition of the three protein kinase configurations to visualize the type of conformational movement. 

In an early study of lysozyme ([McCammon et al. 1976]), the two domains of this protein were assumed to be rigid, and the hinge-bending motion in the presence of solvent was described by the Langevin equation for a damped harmonic oscillator. The angular displacement 0 from the equilibrium position is thus governed by... 

The effective moment of inertia / and the friction coefficient / could easily be estimated. The force constant k associated with the relative motion of the lobes was determined from an empirical energy function. To do so, the molecule was opened in a step-wise fashion by manipulating the hinge region and each resulting structure was energy minimized. Then, the interaction energy between the two domains was measured, and plotted versus 0. 

The catalytic subunit then catalyzes the direct transfer of the 7-phosphate of ATP (visible as small beads at the end of ATP) to its peptide substrate. Catalysis takes place in the cleft between the two domains. Mutual orientation and position of these two lobes can be classified as either closed or open, for a review of the structures and function see e.g. [36]. The presented structure shows a closed conformation. Both the apoenzyme and the binary complex of the porcine C-subunit with di-iodinated inhibitor peptide represent the crystal structure in an open conformation [37] resulting from an overall rotation of the small lobe relative to the large lobe. 

For k = 1, the smoothness class of the boundary dflc can be reduced to Proof. Assume that S is the closed extension of Sc from the class dividing fl into two domains as before. The boundaries dfli,dfl2... 

Extend Sc up to the boundary F such that fl is divided into two domains with Lipschitz boundaries dfli,dfl2- Assume that mens (F n 90 ) > 0, i = 1,2. In each of these domains, for u G the second Korn... 

Let C be a bounded domain with the smooth boundary L, which has an inside smooth curve Lc without self-intersections. We denote flc = fl Tc. Let n = (ni,ri2) be a unit normal vector at L, and n = ( 1,1 2) be a unit normal vector at Lc, which defines a positive and a negative surface of the crack. We assume that there exists a closed continuation S of Lc dividing fl into two domains the domain fl with the outside normal n at S, and the domain 12+ with the outside normal —n at S (see Section 1.4). By doing so, for a smooth function w in flc, we define the traces of w at boundaries 912+ and, in particular, the traces w+ and the jump [w] = w+ — w at Lc. Let us consider the bilinear form... 

Two domains, t1 and t2, exist which affect the GR post-DNA binding transcription activity (37). The major (t1) transactivation domain is 185 amino acid residues ia length with a 58-tesidue a-heUcal functional cote (38). The t1 domain is located at the N terminus of the proteia the minor (t2) trans activation domain residues on the carboxy-terminal side of the DNA binding domain. 

Several motifs usually combine to form compact globular structures, which are called domains. In this book we will use the term tertiary structure as a common term both for the way motifs are arranged into domain structures and for the way a single polypeptide chain folds into one or several domains. In all cases examined so far it has been found that if there is significant amino acid sequence homology in two domains in different proteins, these domains have similar tertiary structures. 

Figure 2.19 Organization of polypeptide chains into domains. Small protein molecules like the epidermal growth factor, EGF, comprise only one domain. Others, like the serine proteinase chymotrypsin, are arranged in two domains that are required to form a functional unit (see Chapter 11). Many of the proteins that are involved in blood coagulation and fibrinolysis, such as urokinase, factor IX, and plasminogen, have long polypeptide chains that comprise different combinations of domains homologous to EGF and serine proteinases and, in addition, calcium-binding domains and Kringle domains.

Serine proteinase domains that are homologous to chymotrypsin, which has about 245 amino acids arranged in two domains. 

Figure 4.21 The polypeptide chain of the arabinose-binding protein in E. coli contains two open twisted a/P domains of similar structure. A schematic diagram of one of these domains is shown in (a). The two domains are oriented such that the carboxy ends of the parallel P strands face each other on opposite sides of a crevice in which the sugar molecule binds, as illustrated in the topology diagram (b). [(a) Adapted from J. Richardson.)...

Figure S.ll A computer-generated diagram of the structure of y crystallin comprising one polypeptide chain of 170 amino acid residues. The diagram illustrates that the polypeptide chain is arranged in two domains (blue and red). Only main chain (N, C , Ca) atoms and no side chains are shown.

Figure S.IS Schematic diagram (a) and topology diagram (b) for the y-crystallin molecule. The two domains of the complete molecule have the same topology each is composed of two Greek key motifs that are joined by a short loop region, [(a) Adapted from T. Blundell et ah. Nature 289 171-777, 1981.]...

A relevant question to ask at this stage is, do the topological identities displayed in the diagram reflect structural similarity We can now see that topologically the polypeptide chain is divided into four consecutive Greek key motifs arranged in two domains. How similar are the domain structures to each other, and how similar are the two motifs within each domain ... 

Tom Blundell has answered these questions by superposing the Ca atoms of the two motifs within a domain with each other and by superposing the Ca atoms of the two domains with each other. As a rule of thumb, when two structures superpose with a mean deviation of less than 2 A they are considered structurally equivalent. For each pair of motifs Blundell found that 40 Ca atoms superpose with a mean distance of 1.4 A. These 40 Ca atoms within each motif are therefore structurally equivalent. Since each motif comprises only 43 or 44 amino acid residues in total, these comparisons show that the structures of the complete motifs are very similar. Not only are the individual motifs similar in stmcture, but they are also pairwise arranged into the two domains in a similar way since superposition of the two domains showed that about 80 Ca atoms of each domain were structurally equivalent. 

Boumann, U., et al. Three-dimensional structure of the alkaline protease of Pseudomonas aeruginosa, a two-domain protein with a calcium binding parallel beta roll motif. EMBO J. 12 3357-3364, 1993. 

Figure 6.6 Schematic diagram of the structure of the enzyme lysozyme which folds into two domains. One domain is essentially a-helical whereas the second domain comprises a three stranded antiparallel p sheet and two a helices. There are three disulfide bonds (green), two in the a-helical domain and one in the second domain.

Figure 6.8 Schematic diagram of the enzyme DsbA which catalyzes disulfide bond formation and rearrangement. The enzyme is folded into two domains, one domain comprising five a helices (green) and a second domain which has a structure similar to the disulfide-containing redox protein thioredoxin (violet). The N-terminal extension (blue) is not present in thioredoxin. (Adapted from J.L. Martin et al.. Nature 365 464-468, 1993.)...

CDK2 has two domains, a small (85 residue) amino-terminal domain comprising a single a helix and a five-stranded p sheet and a larger (213 residues) domain that is mainly a-helical (Figure 6.17a). The cofactor in the... 

Figure 6.21 Schematic diagram of the conformational changes of calmodulin upon peptide binding, (a) In the free form the calmodulin molecule is dumhhell-shaped comprising two domains (red and green), each having two EF hands with bound calcium (yellow), (b) In the form with bound peptides (blue) the a helix linker has been broken, the two ends of the molecule are close together and they form a compact globular complex. The internal structure of each domain is essentially unchanged. The hound peptide binds as an a helix.

Figure 6.21a) comprising two domains separated by a long straight a helix, similar in shape to troponin-C described in Chapter 2 (see Figure 2.13c). Each domain comprises two EF hands (see Figure 2.13a), each of which binds a calcium atom. The two domains are clearly separated in space at the two ends of the a helix linker. 

Each subunit of the homotetrameric PFK of Escherichia coli comprises 320 amino acids arranged in two domains, one large and one smaller, both of which have an rx/p structure reminiscent of the Rossman fold (Figure 6.25). 

Figure 6.25 Schematic diagram of the structure of one dimer of phosphofructokinase. Each polypeptide chain is folded Into two domains (blue and red, and green and brown), each of which has an oi/p structure. Helices are labeled A to M and p strands 1 to 11 from the amino terminus of one polypeptide chain, and respectively from A to M and 1 to 11 for the second polypeptide chain. The binding sites of substrate and effector molecules are schematically marked In gray. The effector site of one subunit is linked to the active site of the other subunit of the dimer through the 6-F loop between helix F and strand 6. (Adapted from T. Schlrmer and P.R. Evans, Nature 343 140-145, 1990.)...

Superficially, the lambda repressor protein is very different from lambda Cro. The polypeptide chain is much larger, 236 amino acids, and is composed of two domains that can be released as separate fragments by mild proteolysis. In repressor the domain responsible for dimerization is separate from the... 

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