No abstract

No abstract

X-ray diffraction studies have produced a low resolution image and also located the iron atoms of a monomeric hemerythrin from muscles of a sipunculan worm. These results reveal the course of the polypeptide chain and some details of the active center.Oxygen transport in certain invertebrate animals is mediated by hemerythrin in erythrocytes of the coelomic fluid. Hemerythrin usually occurs as an octamer of 108,000 molecular weight. It is a non-heme iron protein containing two iron atoms per subunit and it reversibly binds oxygen in the ratio 1 02:2 Fe. Much study has been attended to the structural chemistry of this evolutionary alternative to hemoglobin as an oxygen carrier (1), particularly by Klotz and coworkers, but many details remain obscure. New light can now be shed on hemerythrin structure following the discovery by Klippenstein et al. (2) that the sipunculan worm Themiste (syn. Dendrostomum) pyroides (3) contains a monomeric hemerythrin in its retractor muscles as well as hemerythrin octamers in its erythrocytes-a situation reminiscent of myoglobin and hemoglobin in mammals. Several properties of this myohemerythrin suggest that it bears close structural similarity to the protomers of octameric hemerythrin (2, 4).Myohemerythrin from T. pyroides has been crystallized (4) and the first structural results from studies of these crystals are reported here. These results are mainly at low resolution, but owing to a high helix content and recourse to chemical data, more molecular detail has been gleaned than is ordinarily discernible at low resolution. By way of warning it should be noted that published interpretations of low-resolution density maps have sometimes later been proved incorrect. However, it seems unlikely that such mistakes are repeated here. The quality of this map and the consistency of the model with independent chemical data argue for the basic correctness of the rather detailed molecular interpretation given here.Myohemerythrin is a relatively small protein of 118 aminoacid residues and molecular weight 13,900. This facilitates its crystallographic analysis. In turn, the knowledge of this structure should simplify the analysis of octameric hemerythrins. In any event, further studies on myohemerythrin should be an avenue for gaining a detailed understanding of reversible oxygenation in this fascinating class of proteins. X-ray analysCrystals of metazide myohemerythrin were grown as previously described (4) and then transferred to a stabilizing medium of 80% saturated ammonium sulfate buffered to pH 6.7. These crystals are in space group P2,2121 and have unit cell dimensions of a = 41.58 A; b = 80.03 A; and c = 37.78 X. Derivatives were prepared by soaking native crystals in stabilizing media containing heavy-atom compounds.Many compounds seem to bind, but derivative crystals are often prone to cracking and poor isomorphism. Nevertheless, five derivatives were found to be good enough at least for lowresolution phasing.X-ray diffraction data were measured by w-scans on a fourcircle di...

The complete amino acid sequence of muscle hemerythrin (myohemerythrin) from the sipunculid Themiste (syn. Dendrostomum) pyroides has been determined by analysis of tryptic, chymotryptic, and cyanogen bromide peptides. The primary structure of myohemerythrin differs substantially from that of coelomic hemerythrins of Phascolopsis (syn. Golfingia) gouldii and Themiste pyroides, the amino acid sequence of the muscle protein being only 46 and 45% homologous with the respective coelomic hemerythrins. The most extensive regions of homology between muscle and coelomic proteins occur near the terminii. These and other shorter regions of homology are interpreted in terms of the essential iron ligand residues of the active center.

No abstract

X-ray absorption spectroscopy has been used to study the dimeric iron center in azidomethemerythrin from Phascolopsis gouldii Absorption edge data confirm that the two iron atoms are present as Fe(E) and suggest a hexa-coordination site for each of the iron atoms. The extended x-ray absorption fine structure (EXAFS) analysis provides direct structural evidence of a ,u-oxo bridge between the two iron atoms at an average Fe-O distance of 1. Hemerythrin has been studied by a variety of techniques (1-8) in attempts at elucidating the structure of the active center and understanding the mechanism of reversible oxygenation. Mossbauer, magnetic susceptibility, CD, and optical electronic spectroscopy and x-ray crystallography have provided much detail about the iron environment. In particular, the two irons are antiferromagnetically coupled in oxy-and methemerythrins (7). Cystallographic studies show the coordinating groups to be histidine and carboxylate. Resonance Raman spectroscopy has shown that oxygen is bound as the peroxide (1) and has suggested the presence of OH-, H20, or a ,t-oxo bridge in oxy-and methemerythrin (5).Three crystal structures ofazidomethemerythrins have been determined at the Naval Research Laboratory: monomeric myohemerythrin from Themiste zostericola at 5.5-A resolution (9) with model building and partial refinement to 2.5-A resolution (3), low-resolution structures of octameric hemerythrin from Phascolopsis gouldii (10), and trimeric hemerythrin from Siphonosomafunafuti (unpublished results). The structures of the hydroxylmet and azidomet forms of the octameric hemerythrin from T. dyscrita have been determined and refined to 2.2-A resolution by Stenkamp et aL (11,12) at the University of Washington. Initially, there were substantial disagreements between the two models concerning the Fe-Fe distance and the presence ofa ,u-oxo bridge. More recently, through a series of revisions and refinements at higher resolution, a common model has emerged for the azidomethemerythrin active center. It consists of two hexa-coordinate u-oxo-bridged iron atoms ligated to the protein by five histidyl residues and two bidentate carboxylate bridges.These structures have greatly enhanced our understanding of the hemerythrin active center. Much desired detail is, however, still missing. Electronic information such as the oxidation state is not directly accessible from the crystal structure. Definitive evidence for the presence of a p.-oxo bridge is important, as is verification of the Fe-Fe distance. In addition, structures of the active oxy and deoxy forms (as well as of other met states) have yet to be determined crystallographically. Xray absorption spectroscopy (XAS) has proven useful for investigation of the metal active center in metalloproteins and metalloenzymes (13-15). We have used XAS to examine five derivatives (including the oxy and deoxy) of hemerythrin from P. gouldii and report here the result of our study on the most stable state of the protein, azidomethemerythrin. First, we examine the i...

No abstract

No abstract