Hydroxide Rather Than Histidine Is Coordinated to the Heme in Five-coordinate Ferric Scapharca inaequivalvisHemoglobin

The ferric form of the homodimeric Scapharca hemoglobin undergoes a pH-dependent spin transition of the heme iron. The transition can also be modulated by the presence of salt. From our earlier studies it was shown that three distinct species are populated in the pH range 6-9. At acidic pH, a low-spin six-coordinate structure predominates. At neutral and at alkaline pHs, in addition to a small population of a hexacoordinate high-spin species, a pentacoordinate species is significantly populated. Isotope difference spectra clearly show that the heme group in the latter species has a hydroxide ligand and thereby is not coordinated by the proximal histidine. The stretching frequency of the Fe-OH moiety is 578 cm-1 and shifts to 553 cm-1 in H218O, as would be expected for a Fe-OH unit. On the other hand, the ferrous form of the protein shows substantial stability over a wide pH range. These observations suggest that Scapharca hemoglobin has a unique heme structure that undergoes substantial redox-dependent rearrangements that stabilize the Fe-proximal histidine bond in the functional deoxy form of the protein but not in the ferric form

Oxidized dimeric Scapharca inaequivalvis. Co-driven perturbation of the redox equilibrium.

The dimeric hemoglobin isolated from Scapharca inaequivalvis, HbI, is notable for its highly cooperative oxygen binding and for the unusual proximity of its heme groups. We now report that the oxidized protein, an equilibrium mixture of a dimeric high spin aquomet form and a monomeric low spin hemichrome, binds ferrocyanide tightly which allows for internal electron transfer with the heme iron. Surprisingly, when ferricyanide-oxidized HbI is exposed to CO, its spectrum shifts to that of the ferrous CO derivative. Gasometric removal of CO leads to the oxidized species rather than to ferrous deoxy-HbI. At equilibrium, CO binds with an apparent affinity (p50) of about 10-25 mm of Hg and no cooperativity (20 degrees C, 10-50 mM buffers at pH 6.1). The kinetics of CO binding under pseudo-first order conditions are biphasic (t1/2 of 15-50 s at pH 6.1). The rates depend on protein, but not on CO concentration. The nitrite-oxidized protein is not reduced readily in the presence of CO unless one equivalent of ferrocyanide, but not of ferricyanide, is added. We infer that ferrocyanide, produced in the oxidation reaction, is tightly bound to the protein forming a redox couple with the heme iron. CO shifts the redox equilibrium by acting as a trap for the reduced heme. The equilibrium and kinetic aspects of the process have been accounted for in a reaction scheme where the internal electron transfer reaction is the rate-limiting step

Coordination and spin state equilibria as a function of pH, ionic strength, and protein concentration in oxidized dimeric Scapharca inaequivalvis hemoglobin.

The oxidized homodimeric Scapharca inaequivalvis hemoglobin undergoes changes in coordination and spin state as a function of pH, ionic strength, and protein concentration which have been monitored by optical absorption spectroscopy. Three species contribute to the spectra between pH 5.8 and 8.7: (i) a hexacoordinate high spin aquomet derivative, whose concentration is essentially constant over the whole pH range analyzed; (ii) a pentacoordinate high spin component which prevails at alkaline pH values, and (iii) a hexacoordinate low spin hemichrome, which is formed at acid pH. The contribution of each of the components to the observed spectra was calculated with the singular value decomposition procedure and has been described quantitatively in terms of a linkage scheme which accounts for the change in heme coordination and for the observation that the high spin to low spin transition entails dissociation into monomers. An important feature of the linkage scheme is the cooperative binding of protons to aquomet dimers. Stopped flow experiments to study the kinetics indicate that dissociation into monomers is the rate-limiting process. The unusually strong tendency of oxidized HbI to loose the heme-bound water molecule is discussed in terms of strain in the iron-proximal histidine bond

Long Wavelength VCSELs Exploitation for Low-Cost and Low-Power Consumption Metro and Access Networks

Long wavelength VCSELs are demonstrated to be able to support metro and access networks in order to achieve low-cost and low-power consumption transceivers. In particular, the exploitation of discrete multitone (DMT) direct modulation allows to achieve high transmission capacities and the availability of widely tuneable MEMS-VCSELs to sustain agility, reconfigurability and colourless features of networks

Hagfish Hemoglobins STRUCTURE, FUNCTION, AND OXYGEN-LINKED ASSOCIATION

Cyclostomes, hagfishes and lampreys, contain hemoglobins that are monomeric when oxygenated and polymerize to dimers or tetramers when deoxygenated. The three major hemoglobin components (HbI, HbII, and HbIII) from the hagfish Myxine glutinosa have been characterized and compared with lamprey Petromyzon marinus HbV, whose x-ray crystal structure has been solved in the deoxygenated, dimeric state (Heaslet, H. A., and Royer, W. E., Jr. (1999) Structure 7, 517-526). Of these three, HbII bears the highest sequence similarity to P. marinus HbV. In HbI and HbIII the distal histidine is substituted by a glutamine residue and additional substitutions occur in residues located at the deoxy dimer interface of P. marinus HbV. Infrared spectroscopy of the CO derivatives, used to probe the distal pocket fine structure, brings out a correlation between the CO stretching frequencies and the rates of CO combination. Ultracentrifugation studies show that HbI and HbIII are monomeric in both the oxygenated and deoxygenated states under all conditions studied, whereas deoxy HbII forms dimers at acidic pH values, like P. marinus HbV. Accordingly, the oxygen affinities of HbI and HbIII are independent of pH, whereas HbII displays a Bohr effect below pH 7.2. HbII also forms heterodimers with HbIII and heterotetramers with HbI. The functional counterparts of heteropolymer formation are cooperativity in oxygen binding and the oxygen-linked binding of protons and bicarbonate. The observed effects are explained on the basis of the x-ray structure of P. marinus HbV and the association behavior of site-specific mutants (Qiu, Y., Maillett, D. H., Knapp, J., Olson, J. S., and Riggs, A. F. (2000) J. Biol. Chem. 275, 13517-13528)

Beyond 25 Gb/s Directly-Modulated Widely Tunable VCSEL for Next Generation Access Network

We demonstrate capacities beyond 25Gb/s up to 40 km in the whole C-band range without any dispersion compensation by DMT direct modulation and direct detection exploiting widely tuneable MEMS-VCSELs for future low-cost high-capacity access networks

Immobilization of Lathyrus cicera Amine Oxidase on Magnetic Microparticles for Biocatalytic Applications

Amine oxidases are enzymes belonging to the class of oxidoreductases that are widespread, from bacteria to humans. The amine oxidase from Lathyrus cicera has recently appeared in the landscape of biocatalysis, showing good potential in the green synthesis of aldehydes. This enzyme catalyzes the oxidative deamination of a wide range of primary amines into the corresponding aldehydes but its use as a biocatalyst is challenging due to the possible inactivation that might occur at high product concentrations. Here, we show that the enzyme’s performance can be greatly improved by immobilization on solid supports. The best results are achieved using amino-functionalized magnetic microparticles: the immobilized enzyme retains its activity, greatly improves its thermostability (4 h at 75 ◦C), and can be recycled up to 8 times with a set of aromatic ethylamines. After the last reaction cycle, the overall conversion is about 90% for all tested substrates, with an aldehyde production ranging between 100 and 270 mg depending on the substrate used. As a proof concept, one of the aldehydes thus produced was successfully used for the biomimetic synthesis of a non-natural benzylisoquinoline alkaloid

Selective targeting of melanoma by PEG-masked protein-based multifunctional nanoparticles

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