Single-domain proteins

Larger proteins usually have two or more stmctural units termed domains, each domain having stmctures similar to single-domain proteins. The interaction between individual domains is much less extensive than that within a single domain. In many cases each domain is responsible for carrying out a specific function. 

Plaxco K. W Simons K. T. and Baker D. Contact order, transition state placement and the refolding rates of single domain proteins. J. Mol. Biol. (1998) 277(4) 985-994. 

Figure 13.4 Perturbation of the melting transition temperature of a single domain protein as a function of benzyl alcohol content (described as % B.A.). The liquid protein formulation consists of 10 mM Tris, 4% mannitol, and 1% sucrose, pH 7.4. (Permission to use the figure granted by BioPharm.)...

Ribonuclease H is a small, single domain protein that eleaves RNA from RNA-DNA hybrids. RNase H from E. coli T = 66 °C) has been structurally studied by NMR spectroscopy.Hydrogen exchange NMR experiments have been used to examine the structural distribution of stability in RNase H from T. thermophiliis = 86 °C) and to compare its stability with that of the homologous RNase H from the mesophilic E. The general distribution... 

The following diagram, adapted from that first presented by Bennett et alC, describes a postulated pathway for evolution of a protein dimer from single-domain proteins. The scheme begins with the fusion of two singledomain polypeptides and proceeds through the evolution of interdomain contacts, and in the case of enzymes, development of an active site. These same interdomain contacts can also stabilize formation of a domain-swapped dimer which then undergoes further evolution into a present-day dimer. 

There does, however, appear to be a statistically significant correlation between the rate constants for folding of single domain proteins and the average sequence separation between contacting residues in the native state. Proteins that have primarily local contacts (i.e., have a low contact order) tend to fold more rapidly than those that have more non-local interactions (i.e., have a high contact order).81,82... 

Gelsolin itself was discovered in 1979 by Yin and Stossel based on its ability to activate the gel-sol transformation of actin filaments in a calcium-dependent manner (Yin and Stossel, 1979 Yin et al., 1980). Gelsolin is composed of six domains, named S1-S6, that appears to have arisen from triplication, followed by duplication of an ancestral gene encoding a single domain protein of about 15 kDa (Kwiatkowski et al., 1986) (see Figure 2 Gelsolin structure). 

It has been shown that the hierarchical approach illustrated above for the case of a two-domain protein can be performed at a more fundamental level in order to account for the cooperative folding behavior of single-domain proteins (Freire and Murphy, 1991). This approach involves the use of the crystallographic structure of a pro-... 

Type I copper is present at the active site of blue copper proteins (BCP see chapter by Nersissian and Shipp, this volume) where it is involved in the transfer of a single electron, as well as in multicopper enzymes (Gray et al., 2000 Malmstrdm, 1994 Randall et al., 2000 Sykes, 1991) (see Section V). BCP are single-domain proteins with a (3-barrel fold defined by two (3-sheets that can contain 6 to 13 strands following a Greek-key motif (Fig. 1) (Adman, 1991 Messerschmidt, 1998 Murphy ei a/., 1997 Sykes, 1991). These proteins are stable in both the reduced, Cu(I), and the oxidized, Cu(II), forms. 

Fig. 3.9 (a) A ribbon presentation of the structure of the human tyrosine-specific phosphatase IB, (PTPIB). It is a single-domain protein with a seven-stranded, mixed b-sheet, flanked by a-helices. The catalytic site is in the central region, in a shallow cleft. The phosphate-recognition site is characteristic for PTPs. It is formed by a loop containing the C(X5>R motif, which contains the catalytically essential cysteine and arginine residues. (Reproduced with permission of the authors, D.Barford, A. J. Flint, and N. K.Tonks, and Science from ref. 78.)... 

The catalytic subunit of PPl is a 37 kd single-domain protein. This subunit is usually bound to one of a family of regulatory subunits with masses ol approximately 120 kd in skeletal muscle and heart, the most prevalent regulatory subunit is called Gvr. whereas, in the liver, the most prevalent subunit is Gl. These regulatory subunits have modular structures with domains that participate in interactions with glycogen, with the catalytic subunit, and... 

S. E. Jackson. How do small single-domain proteins fold Folding and Design. 3 (1998), R81. 

The third family of PPIases, the parvulins, does not belong to immunophilins because family members do not show affinity to immunosuppressive drugs. In all cases the prototypic family members are small, single-domain proteins expressed in high abundance predominantly in the cytosol of mammalian and bacterial cells. 

In folded proteins the peptide bonds are usually in the trans conformation, which, for nonprolyl bonds,1 is much less strained than the energetically unfavorable cis conformation. For the peptide bonds that precede proline (prolyl bonds), however, the energy difference between the cis and trans states is small, and therefore cis prolyl bonds are found rather frequently in folded proteins. These cis prolyl bonds create a problem for protein folding. The incorrect trans forms predominate in the unfolded or nascent protein molecules, and the trans —> cis isomer-izations are intrinsically slow reactions because rotation about a partial double bond is required. Incorrect prolyl isomers in a protein chain strongly decelerate its folding. This is clearly seen for small single-domain proteins. Many of them refold within a few milliseconds when they contain correct prolyl isomers but when incorrect isomers are present, folding usually requires seconds to minutes. 

In this section RNase A and RNase T1 are used as examples to illustrate the role of prolyl isomerizations for the unfolding and refolding of small single-domain proteins. Bovine pancreatic RNase A is selected because the history of the proline hypothesis and its experimental verification are closely related with this protein. The mechanism of RNase T1 folding is described because it is one of the major in vitro systems for investigating the function of prolyl isomerases as catalysts of proline-limited protein folding. 

The volume of native Tendamistat is increased by 41.4 + 2.0 cm mol" compared with the denaturated form at pH 2.0 and 35 °C (Fig. 11.2b). This value is virtually independent of the denaturant concentration and it is similar to reaction volumes (AV°) for folding of many other small single-domain proteins [6]. Contributions to AV° may arise from packing deficiencies in the native state as well as from different solvent interactions in the native and in the unfolded protein. Although the reaction volume is virtually independent of the denaturant concentration, the activation volumes for refolding and unfolding both greatly increase with... 

---