The thermal stability of proteins was studied, 195 single amino acid r
esidue replacements reported elsewhere being analysed for several prot
ein conformational characteristics: type of residue replacement; conse
rvative versus nonconservative substitution; replacement being in a ho
mologous stretch of amino acid residues; change in hydrogen bond, van
der Waals and secondary structure propensities; solvent-accessible ver
sus inaccessible replacement; type of secondary structure involved in
the substitution; the physico-chemical characteristics to which the th
ermostability enhancement can be attributed; and the relationship of t
he replacement site to the folding intermediates of the protein, when
known. From the above analyses, some general rules arise which suggest
where amino acid substitutions can be made to enhance protein thermos
tability: substitutions are conservative according to the Dayhoff matr
ix; mainly occur on conserved stretches of residues; preferentially oc
cur on solvent-accessible residues; maintain or enhance the secondary
structure propensity upon substitution; contribute to neutralize the d
ipole moment of the caps of helices and strands; and tend to increase
the number of potential hydrogen bonding or van der Waals contacts or
improve hydrophobic packing.