Letter: Recognition of structural domains in globular proteins

The polypeptide chain of many globular proteins is folded into two or more structural domains which are spatially separated from one another and are frequently based on different architectural principles. The conformation of such structural domains is, in general, conserved more faithfully (assuming divergent evolution) than the amino acid sequence. The preservation of structure may originate in the three-dimensional requirements for the conservation of essential functions. A method is discussed for the recognition and comparison of structural domains.     

Surface activity in vitro: role of surfactant proteins

Pattle, who provided some of the initial direct evidence for the presence of pulmonary surfactant in the lung, was also the first to show surfactant was susceptible to proteases such as trypsin. Pattle concluded surfactant was a lipoprotein. Our group has investigated the roles of the surfactant proteins (SP-) SP-A, SP-B, and SP-C using a captive bubble tensiometer. These studies show that SP-C>SP-B>SP-A in enhancing surfactant lipid adsorption (film formation) to the equilibrium surface tension of approximately 22-25 mN/m from the 70 mN/m of saline at 37 degrees C. In addition to enhancing adsorption, surfactant proteins can stabilize surfactant films so that lateral compression induced through surface area reduction results in the lowering of surface tension (gamma) from approximately 25 mN/m (equilibrium) to values near 0 mN/m. These low tensions, which are required to stabilize alveoli during expiration, are thought to arise through exclusion of fluid phospholipids from the surface monolayer, resulting in an enrichment in the gel phase component dipalmitoylphosphatidylcholine (DPPC). The results are consistent with DPPC enrichment occurring through two mechanisms, selective DPPC adsorption and preferential squeeze-out of fluid components such as unsaturated phosphatidylcholine (PC) and phosphatidylglycerol (PG) from the monolayer. Evidence for selective DPPC adsorption arises from experiments showing that the surface area reductions required to achieve gamma near 0 mN/m with DPPC/PG samples containing SP-B or SP-A plus SP-B films were less than those predicted for a pure squeeze-out mechanism. Surface activity improves during quasi-static or dynamic compression-expansion cycles, indicating the squeeze-out mechanism also occurs. Although SP-C was not as effective as SP-B in promoting selective DPPC adsorption, this protein is more effective in promoting the reinsertion of lipids forced out of the surface monolayer following overcompression at low gamma values. Addition of SP-A to samples containing SP-B but not SP-C limits the increase in gamma(max) during expansion. It is concluded that the surfactant apoproteins possess distinct overlapping functions. SP-B is effective in selective DPPC insertion during monolayer formation and in PG squeeze-out during monolayer compression. SP-A can promote adsorption during film formation, particularly in the presence of SP-B. SP-C appears to have a superior role to SP-B in formation of the surfactant reservoir and in reinsertion of collapse phase lipids.     

Theory for flow of Casson and Herschel-Bulkley fluids in cone-plate viscometers

Mathematical models for blood flow in cone-plate viscometer have been considered, by assuming blood as a Casson/Herschel-Bulkley fluid. Three different cases have been analyzed (i) when there is no shearing, (ii) partial shearing and (iii) full shearing. The relationships between the angular velocity and torque have been obtained for the above three cases. By assuming total shearing, the analytical expression for apparent viscosity has been obtained. Variation of apparent viscosity with yield stress, angular velocity, Casson co-efficient of viscosity, consistency index and flow behaviour index has been computed. It is observed that as the angular velocity increases, the apparent viscosity decreases for both fluids. Further, it is found that as the cone angle increases, the apparent viscosity increases. This behaviour of apparent viscosity in cone-plate viscometer is interesting and unexpected and is being reported first time.     

CASTp: Computed Atlas of Surface Topography of proteins

Computed Atlas of Surface Topography of proteins (CASTp) provides an online resource for locating, delineating and measuring concave surface regions on three-dimensional structures of proteins. These include pockets located on protein surfaces and voids buried in the interior of proteins. The measurement includes the area and volume of pocket or void by solvent accessible surface model (Richards' surface) and by molecular surface model (Connolly's surface), all calculated analytically. CASTp can be used to study surface features and functional regions of proteins. CASTp includes a graphical user interface, flexible interactive visualization, as well as on-the-fly calculation for user uploaded structures. CASTp is updated daily and can be accessed at http://cast.engr.uic.edu.     

Surface beta-strands in proteins: identification using an hydropathy technique

From a representative set of monomeric globular proteins with known three-dimensional structures, beta-strands with lengths > or = 5 amino acids have been identified and catalogued. By ascertaining the accessible surface areas of the constituent residues in these strands, and by checking whether the exposed/buried pattern is 80% or more similar to that in an idealized surface strand, a subset of structures can be delineated in which the beta-strands are all sited on the surface of the protein. The corresponding sequence data show that about 50% of the residues are apolar (Val, Ile, Leu, Phe, Tyr, Ala) and that the common occurrence of valine (14.3%), isoleucine (9.6%), and threonine (8.1%) is a characteristic feature. The frequencies of occurrence of those amino acids in the strands that face the aqueous environment and the interior have also been determined separately and show that most surface strands have a substructure of the form (apolar-X)(n), where X is approximately equally divided between apolar, charged, and hydrophilic residues. Using the frequency data thus obtained, allied with an algorithm to delineate potential surface beta-strands from characteristic hydropathy profiles, it is now possible to search through the sequences of proteins with unknown tertiary structures and make realistic predictions of the presence of this element of structure on the protein surface. In addition, new data are presented on the distribution of the various types of residues on the surface of proteins and in their interior. Significant differences were observed, not all of which have been identified previously. Furthermore, the distribution of the types of residue in a surface beta-strand was compared to that corresponding to the surfaces of all of the proteins in our database. Again, very characteristic differences were observed. These are helpful in recognizing the presence of surface beta-strands.     

Surface shear viscometry as a probe of protein-protein interactions in mixed milk protein films adsorbed at the oil-water interface

Time-dependent surface viscosities are reported for films adsorbed from binary mixtures of the proteins alpha-lactalbumin, beta-lactoglobulin and beta-casein. The measurements were made at a planar interface between n-tetradecane and various protein solutions (10(-3) wt% of each protein, pH 7, 25 degrees C) using a Couette-type torsion-wire surface viscometer operating at very low shear-rate. Differences in behaviour between simultaneous and sequential exposure of the pairs of proteins to the interface were investigated. Some experiments were performed with chemically modified beta-lactoglobulin samples whose disulphide bonds had been cleaved and blocked. Displacement of one protein by another (e.g. alpha-lactalbumin by beta-casein) is indicated by a sudden drop in surface viscosity immediately after addition of the second protein. In systems containing beta-lactoglobulin, the long-time surface viscosity is very sensitive to the adsorption time of beta-lactoglobulin prior to addition of the second protein. Blocking the disulphide bonds in beta-lactoglobulin leads to a much faster approach to a steady-state surface viscosity. This is interpreted in terms of a much more rapid unfolding of the disordered molecules of modified beta-lactoglobulin at the oil-water interface. We conclude that surface viscosity experiments give useful and sensitive information about competitive adsorption and cooperative interactions in mixed protein films.     

Determination of the Surface tension of proteins. I. Surface tension of native serum proteins in aqueous media

The desorption patterns of serum proteins in hydrophobic chromatography suggest that serum proteins that remain immersed in an aqueous medium and do not become in a protein-air interface are very hydrophilic. Contact angle measurements on fairly thick layers of hydrated serum proteins, formed on ultrafiltration membranes, yield surface tensions that correlate well with the degree of hydrophilicity derived from desorption data obtained by hydrophobic chromatography. For further confirmation the absorptivity of four human serum proteins was measured with respect to surfaces of different polymers of various surface tensions, for solution in aqueous solvents of different surface tensions. The surface tension of the solvent from which a dissolved protein adsorbs to precisely the same extent onto all solid substrates (regardless of their surface tensions) is equal to the surface tension of that protein. The surface tensions found by the contact angle (first value given) and by the protein adsorption methods (second value given) were. in erg/cm2; alpha 2-macroglobulin, 71.0, 71.0; serum albumin, 70.5, 70.2; immunoglobulin M, 69.5, 69.4; immunoglobulin G, 67.4, 67.7.     

Surface map comparison: studying function diversity of homologous proteins

A simplified protein surface cartography approach has been developed to assist in the analysis of surface features in homologous families, and thus to predict conservation or divergence of protein functions and protein-protein interaction patterns. A spherical approximation of protein surface was used, with a focus on charged and hydrophobic residues. The resulting surface map allows for qualitative analysis and comparison of surfaces of proteins, but can also be used to define a simple numerical measure of map similarity between two or more proteins. The latter was shown to be useful for function based classifications within large protein families.Surface map analysis was tested on several test cases: haemoglobins, death domains and TRAF domains. It was shown that surface map comparison allows a better function prediction than general sequence analysis methods and can reproduce known examples of functional variation within a divergent group of proteins. In another example, we predict novel, unexpected sets of common functional properties for seemingly distant members of a large group of divergent proteins. The method was also shown to be robust enough to allow using protein models from comparative modelling instead of experimental structures.