Reduced proteins, thermostability

We will discuss three different approaches to engineer a more thermostable protein than wild-type T4 lysozyme, namely (1) reducing the difference in entropy between folded and unfolded protein, which in practice means reducing the number of conformations in the unfolded state, (2) stabilizing tbe a helices, and (3) increasing the number of bydropbobic interactions in tbe interior core. 

The term thermal stability (also thermostability) refers to the resistance of a protein to adverse intrinsic and extrinsic environmental influences, i.e., the thermal characteristic of the protein to remain steady against the dena-turation of its molecular integrity and inactivation of its biologic activity on facing high temperatures or other deleterious agents (6). One of the most important indices to measure protein stability is the decimal reduction time, or D-value, the time required to reduce 90% of the initial protein concentration exposed to the reference temperature. The D-value was used... 

Biopharmaceuticals may be subjected to heat treatment as a means of viral inactivation. Protein damage is prevented by the addition of high concentrations of a thermostabilizing excipient such as sugars or polyhyd-ric alcohols. The non-reducing polyhydric additives increase thermostability of the protein by the formation of a hydrogen-bonded solvent shell. The typical treatment for factor VIII consists of heat treatment at 60°C for... 

Thermostability of Drifted Oligomeric, Reduced, and Refolded Proteins... 

Upon addition of proteins to aprotic, hydrophobic solvents, the increased intramolecular interactions of the lyophilized protein result in restricted conformational mobility of the protein, thus restricting activity [30], However, proteins display increased thermostability in anhydrous organic solvents as a result of the reduced conformational mobility [31], and water-free methods may help avoid aggregation processes that occur when using the double-emulsion technique, in which the protein is conformationally mobile. 

Fund., Genet., 13, 87 (1992). Mutating the Charged Residues in the Binding Pocket of Cellular Retinoic Acid-Binding Protein Simultaneously Reduces Its Binding Affinity to Retinoic Acid and Increases Its Thermostability. 

Poly(ethylene glycol) (PEG) has been used to modify a number of therapeutically interesting proteins. It has been clearly demonstrated by Abuchowski et al. that grafting of PEG onto proteins reduces their immunogenicity, improves their resistance to proteolytic degradation and improves their thermostability. 

Figure 2. Solution thermostability of rPST and CM-PST. Samples were initially dissolved at lOOmg/ml in phosphate-buffered saline and incubated at 43°C. The percentage of the original protein that remained in the soluble monomeric form is shown by bars for normal recombinant PST and chemically modified PST, which was reduced and alkylated at the small-loop disulfide. The amount of dimerization of these species as a percentage of the protein remaining in solutions is also shown. (Data from Buckwalter et a ., 1992.)...

SOD and other antioxidant enzymes are produced commercially on a limited scale, mainly for laboratory purposes, being isolated from red blood cells. Propionibacteria are a good source of SOD and their value in this respect rises due to the possibility of multi-purpose processing of the biomass. We have developed (Kraeva and Vorobjeva, 1981a, b) a simple method of isolation and purification of SOD to apparent homogeneity, which is shown in Table 7.5. Since SOD is a thermostable protein, heat treatment was used in the purification, thus significantly reducing the number of purification steps. In the course of purification no enzyme modification was... 

The low molecular weight subfraction of protein complex has mitosis-inhibitory properties, reduces number of active and moderately active nucleoli, and decreases the activation of RNA synthesis in nuclei of car-diomiocytes of newborn rat [12, 13]. Usually, this subfiaction is essential component of any thermostable protein complex obtained from various organs of adult rat. It is always seen as a major subfraction in native gel-electrophoresis (PAAG electrophoresis) (Figure 19.3). 

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