Structural and thermodynamic consequences of b heme binding for monomeric apoglobins and other apoproteins

Abstract

The binding of a cofactor to a protein matrix often involves a reorganization of the polypeptide structure. b Hemoproteins provide multiple examples of this behavior. In this minireview, selected monomeric and single b heme proteins endowed with distinct topological properties are inspected for the extent of induced refolding upon heme binding. To complement the data reported in the literature, original results are presented on a two-on-two globin of cyanobacterial origin (Synechococcus sp. PCC 7002 GlbN) and on the heme-containing module of FixL, an oxygen-sensing protein with the mixed alpha/beta topology of PAS domains. GlbN had a stable apoprotein that was further stabilized and locally refolded by heme binding; in contrast, apoFixLH presented features of a molten globule. Sequence analyses (helicity, disorder, and polarity) and solvent accessibility calculations were performed to identify trends in the architecture of b hemoproteins. In several cases, the primary structure appeared biased toward a partially disordered binding pocket in the absence of the cofactor.

Structural propensities in the heme binding region of apocytochrome b5. I. Free peptides

“…Its hydrophobicity allows it to form strong nonspecific interactions with nonpolar side chains, which, added to specific interactions and ligation of the central iron, can shape the binding site and have a profound influence on holoprotein structure. 9 For some hemoproteins, all or part of the polypeptide chain remains essentially unstructured in the apoprotein state. A close examination of such regions, however, suggests that holo-and nonholo-like structural Structural Propensities in the Heme Binding Region of Apocytochrome b 5 .…”

“…14 It is noteworthy that helices H3 and H4 are identified by the programs IUPred 15 and VSL2, 16,17 as elements of the protein with a tendency to be unstructured. 9 In both apo-and holoprotein solution structures, the terminal backbone atoms of the heme-binding region (Leu32 N and Thr73 C) are held *12 Å apart by the folded protein scaffold (see Figure 1). Restraining the ends of a 42-residue Gaussian chain entails a large entropic penalty, estimated to contribute a Gibbs energy of *24 kJ/mol against the native state at room temperature.…”

Structural propensities in the heme binding region of apocytochrome b5. II. Heme conjugates

“…In some cases, the location of the affected regions can be predicted from trends in the primary structure. 1 The residues involved tend to be polar and, in long stretches, favor disordered conformations. 2 Intrinsically unfolded regions of sequence appear to be essential features in all proteomes.…”

Structural and thermodynamic encoding in the sequence of rat microsomal cytochrome b5

“…25 A similar analysis of multiple b heme protein sequences reveals a trend for regions of low folding propensity near the heme group. 26 The energetics of the cytochrome can be explored with amino acid replacements. For our purposes, these can be organized in two types.…”