Hemopexin structure

Wu, M. L. Interaction of heme with hemopexin Structural characterization and functional imphcations Rh.D. Dissertation, University of Missouri—Kansas City, 1994, pp. 1-156. 

Figure 2 Domain structure of the MMPs 92 kDa gelatinase-A (MMP-2), 72 kDa gelatinase-B (MMP-9), the collagenases (MMP-1, -8, and -13), stromelysin-1 (MMP-3) and matrilysin (MMP-7). Matrilysin is the only known MMP that does not have a C-terminal hemopexin-like domain.

Bode W. A helping hand for collagenases the hemopexin-like domain. Structure 1995 3 527-530. 

Here, the structure and function of hemopexin, the mechanisms of hemopexin-mediated heme transport, and the biological consequences of this specific transport system are reviewed and questions for future research proposed. This is an opportune time for review since recent advances not only provide new viewpoints and directions but also enable new insights to be derived from earlier work. 

Binding of heme by isolated N-domain causes a change in sedimentation coefficient consistent with a more compact conformation and leads to the more avid association with the C-domain (125). Sedimentation equilibrium analysis showed that the Kd decreases from 55 pM to 0.8 pM (Fig. 5) (106). In addition, the calorimetric AH (-1-11 kcal/mol) and AS (-1-65 kcal/mol K) for the heme-N-domain-C-domain interaction and the AH (-3.6 kcal/mol) and AS (-1-8.1 kcal/mol K) derived from van t Hoff analysis of ultracentrifuge data for the interaction in the absence of heme indicate that hydrophobic interactions predominate in the presence of heme and a mix (e.g., hydrophobic and van der Waals forces) drives the interaction in the absence of heme. However, FTIR spectra (Fig. 6) indicate that little change in the secondary structure of domains or intact hemopexin occurs upon heme binding (104). 

Pig. 4. CD spectra in the near and far UV of apo- and heme-hemopexin. The CD spectra of rabbit apo- and heme-hemopexin (solid line and dashed line, respectively) at pH 7.4 in 0.05 M sodium phosphate buffer are shown. The increases in ellipticity in the near UV are attributable to changes in tertiary conformation leading to altered environments of aromatic residues, particularly tryptophan. The unusual positive ellipticity in the far UV is attributable to tryptophan-tryptophan interactions that are perturbed by heme binding 124, 130). This positive signal precludes analysis of the secondary structure of hemopexin using current CD-based algorithms. 

The structure of the C-domain of hemopexin was determined first (128). The structure is a four-hladed p-propeller (Fig. 7), the smallest P-propeller known, and serves as the paradigm for the several proteins known to have a pexin domain, including vitronectin (108), and several metalloproteinases (107). The repeats evident in the sequence of hemopexin (99-101), for instance DAAV/F motifs and WD repeat, form a large part of the p-strands of the four blades, which are connected by short loops and a-helices. 

This structure looks stable because of the compact, symmetric structure with multiple reinforcing interactions, and direct examination of hemopexin (vide infra) has home this out. Smith et al. (129) have... 

Fig. 6. Deconvolved amide F region FTIR spectra of apo- and heme-hemopexin. The amide F FTIR spectra of apo- and heme-hemopexin in D2 O were recorded and curve-fitted to resolve the individual bands. The differences between the original and fitted curves are shown in the upper traces in the panels. The estimated helix (15%), beta (54%), turn (19%), and coil (12%) content of the apo-protein are not significantly changed upon heme binding 104). This analysis was required because of the positive 231-nm elhpticity band in hemopexin and is consistent with the derived crystal structure results.

Fig. 7. The crystal structure of the C-domain of hemopexin (PDB accession number IHXN) 128) showed a four-bladed p-propeller structure, which because of sequence similarity was also expected in the N-domain. The high degree of beta structure and limited a-helix content agrees with the earlier FTIR analysis.

Determination of the structure of the entire rabbit heme-hemopexin complex (11) (Fig. 8) clearly revealed the expected complementary structures of the N- and C-domains. More importantly, several fundamental... 

Fig. 8. Crystal structure of heme-hemopexin. The crystal structure of the rabbit mesoheme-hemopexin complex (PDB accession number IQHU) (11) showed heme to be bound in a relatively exposed site between the N- and C-domains with one axial His ligand being contributed by the hinge or linking region between the domains and the other by the C-domain. Also noteworthy is the disposition of the heme with its propionate residues pointing inward and neutralized by positive charges in the binding site.

Fig. 15. Effects of pH on apo- and heme-hemopexin. The Soret region absorbance (filled squares) of rabbit heme-hemopexin was monitored in two separate titrations, from pH 7.4 to 11.8 in one and from pH 7.4 to 3.8 in the other. Similarly, theellipticity at 231 nm of apo-hemopexin (open circles) and of heme-hemopexin (filled circles) was assessed from pH 7.4 to 11.8 and from pH 7.4 to 1.7 111). The heme complex and the tertiary structure are unaffected by pH in the region from pH 6 to 9, and other values are normalized to these.

A working model of the mechanism of hemopexin-mediated heme uptake (Fig. 17) has been formulated that not only incorporates the known facts concerning the structure of hemopexin, the interaction of hemopexin with heme and its receptor, the factors that influence these interactions, and the cellular responses to hemopexin, but also postulates additional features likely to be needed for the system to function efficiently. As such this scheme represents a testable hypothesis and basis for future study of the hemopexin system. 

A) as the catalytic-domain structure of HFC. This demonstrates that the absence or presence of the hemopexin domain does not affect the overall structure of the catalytic domain. 

Figure 14 (a) The X-ray structure of heme-bound rabbit hemopexin. (b) shows the /S-propeller structure of individual domains. The figure was created using pdb coordinates Iqhu ... 

To date, the crystal structures of 12 different MMPs have been solved. Full structures were obtained for MMP-1 (2CLT), MMP-2 (1CK7), and MMP-7 (IMMP). As for the rest, only the catalytic domains in the presence of different inhibitors were determined. The hemopexin-like domains of MMP-2 (IRTG), MMP-9 (IITV), and MMP-13 (IPEX) were crystallized, and structures were determined separately. Nuclear magnetic resonance (NMR) structures of the catalytic domains of MMP-1 (1AYK), MMP-2 (IHOV), MMP-3 (lUMS), MMP-12 (1YCM), and MMP-13 (lEUB) have also become available. 

The chemistry and biochemistry of Hpx has been reviewed and a crystal structure is available. Hemopexin is present in serum at about 10 pM and its primary function is to transport released heme to its degradation site in the parenchymal cells of the liver via receptor-mediated endocytosis. Encapsulation of a single heme by Hpx occurs via bis-histidyl protein side-chain coordination of the Fe. Spectroelectrochemical investigation of the heme-Hpx assembly gives insight into the role of Hpx in controlling the reduction potential of the heme Fe, the efficiency of electron transfer at the metal centre, the influence of bis-histidyl coordination at the Fe centre, and the possible role of Fe redox in the Hpx-mediated transport and recycling of heme. 

The redox sensitivity of hemopexin-encapsulated heme to electrolyte composition and pH illustrate the importance of first coordination shell (bis-histidine ligation and heme structure) and second coordination shell (protein structure/folding and environment) effects in these heme proteins. These observations also suggest a possible role for Fe " /Fe redox in hemopexin-mediated heme transport/recycling, as high chloride anion concentration and low pH are known conditions for the endosome where the heme is released. 

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