Arginine residues as stabilizing elements in proteins

Abstract

Site-specific substitutions of arginine for lysine in the thermostable D-xylose isomerase (XI) from Actinoplanes missouriensis are shown to impart significant heat stability enhancement in the presence of sugar substrates most probably by interfering with nonenzymatic glycation. The same substitutions are also found to increase heat stability in the absence of any sugar derivatives, where a mechanism based on prevention of glycation can no longer be invoked. This rather conservative substitution is moreover shown to improve thermostability in two other structurally unrelated proteins, human copper, zinc-superoxide dismutase (CuZnSOD) and D-glyceraldehyde-3-phosphate dehydrogenase (GAPDH) from Bacillus subtilis. The stabilizing effect of Lys----Arg substitutions is rationalized on the basis of a detailed analysis of the crystal structures of wild-type XI and of engineered variants with Lys----Arg substitution at four distinct locations, residues 253, 309, 319, and 323. Molecular model building analysis of the structures of wild-type and mutant CuZnSOD (K9R) and GAPDH (G281K and G281R) is used to explain the observed stability enhancement in these proteins. In addition to demonstrating that even thermostable proteins can lend themselves to further stability improvement, our findings provide direct evidence that arginine residues are important stabilizing elements in proteins. Moreover, the stabilizing role of electrostatic interactions, particularly between subunits in oligomeric proteins, is documented.

Stability and structural analysis of α‐amylase from the antarctic psychrophile Alteromonas haloplanctis A23

“…A. haloplanctis a-amylase also possesses a low arginine content. Since the charge resonance of the guanidinium group gives arginine the possibility to form more than one electrostatic bond (Mrabet et al, 1992), an increased arginine content has been correlated with heat stability in thermophilic enzymes (Merkler et al, 1981). The disulfide bond Cys70-CysllS is absent in A. haloplanctis a-amylase.…”

The cloning, expression and crystallisation of a thermostable arginase

“…Cysteine, asparagine and glutamine are thought to be susceptible to covalent damage at high temperatures (e.g. deamidation) and thus tend to be replaced in thermostable proteins [26]. A comparison of the amino acid sequence of the arginase from 'B.…”

Unique molecular properties of superoxide dismutase from teleost fish skin

“…Therefore, one of the reasons for the low thermal stability of SOD-Fs, seems to be concerned with low amounts of hydrophobic AAs, high amounts of polar AAs and the strength of non-covalent interactions between the subunits of SOD-Fs. Non-covalent interactions, such as electrostatic interactions and hydrogen bonds, are also thought to contribute to protein stability and flexibility when they occur on the surface of proteins [37,38]. Furthermore, it is also pointed out .…”