Quaternary structure of the giant extracellular hemoglobin of the leech Macrobdella decora

Kapp, Oscar H.; Qabar, Aziz N.; Bonner, Michael; Stern, Mary S.; Walz, Daniel A.; Schmuck, Maria; Pilz, Ingrid; Wall, Joseph S.; Vinogradov, Serge N.

doi:10.1016/s0022-2836(05)80127-5

Cited by 43 publications

(19 citation statements)

References 41 publications

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“…The Hb was prepared as described earlier for the leech Macrobdella decora, by rapid homogenization in the cold room, in 0.1 M Tris.Cl buffer, 1 mM EDTA at neutral pH in the presence of a protease inhibitor cocktail (no. 1697-498, Biochemical Division, Boehringer Mannheim Corp., Indianapolis, IN) [10]. The homogenate was immediately centrifuged at ϳ20,000 ϫ g for 20 min at 4°C to remove cellular debris; this step was repeated as many times as necessary until a clear solution was obtained.…”

Section: Methodsmentioning

confidence: 99%

An electrospray ionization mass spectrometric study of the subunit structure of the giant hemoglobin from the leech Nephelopsis oscura

Green

Vinogradov

2004

J. Am. Soc. Mass Spectrom.

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The subunit structure of the giant, extracellular hexagonal bilayer (HBL) hemoglobin (Hb) from the leech Nephelopsis oscura was investigated by electrospray ionization mass spectrometry (ESI-MS) employing a maximum entropy deconvolution of its complex, multiply charged ESI spectra. The denatured unreduced Hb consisted of three monomer globin chains These giant complexes were found to have masses of ϳ3500 kDa and to consist of two types of polypeptide chains: heme-containing 16 -17 kDa globin chains and non-globin, linker chains of 24 -32 kDa containing 9 to 12 cysteines [4]. The amino acid sequences of the globin chains are clearly related to vertebrate and invertebrate globins [5] and they form disulfide-bonded dimer, trimer and tetramer subunits as well as noncovalent subassemblies, ranging from dimer to dodecamer [4].Over the last decade, ESI-MS has been shown to be the method of choice for the characterization of proteins [6,7] by providing highly accurate masses. In particular, in the case of heteromultimeric protein complexes, as exemplified by the HBL Hbs, ESI-MS provides an exquisitely accurate enumeration of the constituent chains and covalently bonded subunits, as well as a quantitative determination of free and disulfidebonded Cys residues [8]. Furthermore, the combination of nanoflow electrospray and time-of-flight (TOF) mass analyzers has recently allowed the investigation of the quaternary structure of large noncovalent protein complexes [9].We describe here the results of ESI-MS and ESI-TOF-MS studies of the polypeptide chain and subunit composition and noncovalent subassemblies of the HBL Hb from the leech Nephelopsis oscura.

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Section: Methodsmentioning

confidence: 99%

An electrospray ionization mass spectrometric study of the subunit structure of the giant hemoglobin from the leech Nephelopsis oscura

Green

Vinogradov

2004

J. Am. Soc. Mass Spectrom.

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“…The presence of a dodecamer subassembly has been demonstrated in Eudistylia Chl (441) but not in the Hb of the leech Macrobdella decora (290). The latest three-dimensional reconstruction of Lumbricus Hb at 2.2-nm resolution by Taveau et al (538) revealed all 144 globin chains organized into 12 dodecamer subassemblies and 42 linker subunits.…”

Section: Giant Hbl Hbs and Chls: Summit Of Hb Complexitymentioning

confidence: 99%

Nonvertebrate Hemoglobins: Functions and Molecular Adaptations

Weber

Vinogradov

2001

Physiological Reviews

452

404

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Hemoglobin (Hb) occurs in all the kingdoms of living organisms. Its distribution is episodic among the nonvertebrate groups in contrast to vertebrates. Nonvertebrate Hbs range from single-chain globins found in bacteria, algae, protozoa, and plants to large, multisubunit, multidomain Hbs found in nematodes, molluscs and crustaceans, and the giant annelid and vestimentiferan Hbs comprised of globin and nonglobin subunits. Chimeric hemoglobins have been found recently in bacteria and fungi. Hb occurs intracellularly in specific tissues and in circulating red blood cells (RBCs) and freely dissolved in various body fluids. In addition to transporting and storing O(2) and facilitating its diffusion, several novel Hb functions have emerged, including control of nitric oxide (NO) levels in microorganisms, use of NO to control the level of O(2) in nematodes, binding and transport of sulfide in endosymbiont-harboring species and protection against sulfide, scavenging of O(2 )in symbiotic leguminous plants, O(2 )sensing in bacteria and archaebacteria, and dehaloperoxidase activity useful in detoxification of chlorinated materials. This review focuses on the extensive variation in the functional properties of nonvertebrate Hbs, their O(2 )binding affinities, their homotropic interactions (cooperativity), and the sensitivities of these parameters to temperature and heterotropic effectors such as protons and cations. Whenever possible, it attempts to relate the ligand binding properties to the known molecular structures. The divergent and convergent evolutionary trends evident in the structures and functions of nonvertebrate Hbs appear to be adaptive in extending the inhabitable environment available to Hb-containing organisms.

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“…These proteins exhibit variable heterotropic and homotropic interactions: Bohr effects and cooperativity between the heme groups, respectively, that modulate oxygen transport for a given difference in oxygen tensions at the sites of oxygen loading (gills) and unloading (metabolizing tissues). They consist of hexagonal bilayer (HBL) 1 molecules composed of two superimposed rings that each contain six dodecamers, each comprising three tetramers (12 oxygen-binding heme groups) as well as a number of heme-free "linker" chains that tether the structure (1)(2)(3)(4)(5). The hierarchical arrangement of the leech hemoglobin can thus be written as 2 ϫ (6 ϫ (3 ϫ 4)).…”

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confidence: 99%

“…In M. decora Hb that dissociates into tetrameric, dimeric, and monomeric units and few, if any, dodecamers (2,8), the intact molecules and the tetrameric and monomeric subunits show increasing affinities (oxygen partial pressures at halfsaturation (p 50 ) of 4.4, 1.9, and 0.3 torr, respectively, at pH 7.5 and 25°C). Furthermore, decreasing Bohr factors ( ϭ Ϫ0.38, Ϫ0.30, and 0, respectively) and Hill coefficients at half-saturation (n 50 ϭ 3.1, 1.4, and 1.0, respectively) were observed (8).…”

mentioning

confidence: 99%

Nested Allosteric Interactions in Extracellular Hemoglobin of the Leech Macrobdella decora

Hellmann

Weber

Decker

2003

Journal of Biological Chemistry

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Hemoglobin from the leech Macrobdella decora belongs to the class of giant extracellular hexagonal bilayer globin structures found in annelid and vestimentiferan worms. These complexes consist of 144 hemebearing subunits, exhibit a characteristic quaternary structure (2 ؋ (6 ؋ (3 ؋ 4))), and contain tetramers as basic substructures that express cooperative oxygen binding and thus provide a structural basis for a hierarchy in allosteric interactions. A thorough analysis of the isolated tetramer indicates that it functions as a trimer of cooperatively interacting subunits and a noncooperative monomer rather than as four interacting subunits. A thermodynamic analysis of the whole molecule favors the application of a nested Monod-WymanChangeux model with six cooperatively interacting 12-mer allosteric units. In contrast to the isolated tetramers, all subunits of the tetramers seem to be coupled cooperatively within the oligomerized 144-mer. Thus, besides hemocyanins and GroEL, the hexagonal bilayer hemoglobins represent another class of proteins in which the hierarchical quaternary structure provides the basis for nested interaction in their functional properties.In contrast to vertebrate animals that possess tetrameric hemoglobins in red blood cells and monomeric myoglobin in muscle cells, many invertebrates have extracellular hemoglobins with enormously high molecular weights. Based on electron microscopic and structural analyses, the extracellular hemoglobins of annelids exhibit unique structures that constitute the summit of complexity for oxygen-binding hemes. These proteins exhibit variable heterotropic and homotropic interactions: Bohr effects and cooperativity between the heme groups, respectively, that modulate oxygen transport for a given difference in oxygen tensions at the sites of oxygen loading (gills) and unloading (metabolizing tissues). They consist of hexagonal bilayer (HBL) 1 molecules composed of two superimposed rings that each contain six dodecamers, each comprising three tetramers (12 oxygen-binding heme groups) as well as a number of heme-free "linker" chains that tether the structure (1-5). The hierarchical arrangement of the leech hemoglobin can thus be written as 2 ϫ (6 ϫ (3 ϫ 4)). This quaternary structure is supported by a recent x-ray structure resolution of the HBL hemoglobin from the earthworm Lumbricus terrestris (6), which similarly consists of two superimposed six-membered rings of dodecamers and can be dissociated into trimeric and monomeric subunits (7). Cooperativity between the heme groups of leech Macrobdella decora Hb is low (maximum Hill coefficient (n H ) of ϳ3) compared with the large number of heme groups, and both the cooperativity coefficient and the oxygen affinity (Bohr factor ( ) of Ϫ0.4) are pH-sensitive (8,9).No conclusive information is available with regard to the influence of hierarchical structure on the expression of cooperative and allosteric interactions in annelid hemoglobins. This is in part because of differences in the functional criteria considered in d...

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Quaternary structure of the giant extracellular hemoglobin of the leech Macrobdella decora

Cited by 43 publications

References 41 publications

An electrospray ionization mass spectrometric study of the subunit structure of the giant hemoglobin from the leech Nephelopsis oscura

An electrospray ionization mass spectrometric study of the subunit structure of the giant hemoglobin from the leech Nephelopsis oscura

Nonvertebrate Hemoglobins: Functions and Molecular Adaptations

Nested Allosteric Interactions in Extracellular Hemoglobin of the Leech Macrobdella decora

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