Non-functional conserved residues in globins and their possible role as a folding nucleus

Structure-based sequence alignment of 728 sequences of different globin sub families shows that in each subfamily there are two clusters of consensuall y conserved residues. The first is the well-known "functional" cluster whic h includes six heme-binding conserved residues (Phe CD1, His F8; aliphatic Ell, FG5; hydrophobic F4, G5) and seven other conserved residues (Pro C2; a liphatic H19; hydrophobic B10, B13, B14, CD4, E4) that do not bind the heme but belong to its immediate neighborhood. The second cluster revealed here (aliphatic A8, G16, G12; aromatic A12; hydrophobic H8 and possibly H12) is distant from the heme. It is entirely non-polar and includes one turn (i, i+4 positions) from each of helices A, G, and H. It is known that A, G, and H helices formed at the earliest stage of apomyoglobin folding remain rela tively stable in the equilibrium molten globule state, and are likely to be tightly packed with each other in this state. We have shown the existence of two similar conserved clusters in c-type cytochromes, heme-binding and d istal from the heme. The second cluster in c-cytochromes includes one turn from each of the N and C-terminal a-helices. These N and C-terminal helices in cytochrome c are formed at the earliest stage of protein folding, remai n relatively stable in the molten globule state, and are tightly packed wit h each other in this state, similar to the observed behavior of the globins . At least these two large protein families (c-type cytochromes and globins ) have a close similarity in the existence and mutual positions of non-func tional conserved residues. We assume that non-functional conserved residues are requisite for the fast and correct folding of both of these protein fa milies into their stable 3D structures.