Thermodynamics of oxygenation-linked proton and lactate binding govern the temperature sensitivity of O2 binding in crustacean(<i>Carcinus maenas</i>) hemocyanin

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SUMMARYThe inverse relationship between temperature and hemoglobin-O 2 affinity resulting from the exothermic nature of heme oxygenation favors O 2 unloading from blood to warm, metabolically active tissues. However, this temperature sensitivity is maladaptive, and commonly countered in regional heterotherms, where it may hamper unloading (e.g. in cold extremities of arctic mammals) or increase the diffusive arterio-venous short-circuiting of O 2 (e.g. in counter-current heat exchangers of warm swimming muscles of tuna). We hypothesized analogous blood specializations in heterothermic billfish, whose warm eyes and brains increase the temporal resolution of vision, and measured hemoglobin-O 2 binding properties in three species over a wide pH range, at two temperatures, and in the absence and presence of the major red cell effector, ATP, permitting detailed assessment of overall oxygenation enthalpies (⌬HЈ) and contributions from oxygenation-linked proton and ATP dissociation. . Thus in addition to allosterically modulating hemoglobin-O 2 affinity, ATP diminishes its temperature sensitivity, reducing deleterious arterio-venous short-circuiting of oxygen in the cranial billfish heat exchangers. The mechanism underlying this reduction in oxygenation enthalpy differs fundamentally from that in tuna, supporting independent evolution of this trait in these scombroid lineages.

Section: Methodsmentioning

confidence: 99%

ATP-induced temperature independence of hemoglobin–O2 affinity in heterothermic billfish

Weber

Campbell

Fago

et al. 2010

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“…Hc concentration was derived from the absorbance at 335nm using the ε=0.0175lmol -1 cm −1 reported for crab Carcinus manaes Hc (Nickerson and Van Holde, 1971;Weber et al, 2008) and a subunit mass of 73.5kDa as deduced from cDNA sequence analysis (described below).…”

Section: Animals and Hemolymphmentioning

confidence: 99%

Molecular and functional characterization of hemocyanin of the giant African millipede,Archispirostreptus gigas

Damsgaard

Fago

Hagner-Holler

et al. 2013

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SUMMARYIn contrast to other terrestrial arthropods, where gaseous O 2 that fuels aerobic metabolism diffuses to the tissues in tracheal tubes, and most other metazoans, where O 2 is transported to tissues by circulating respiratory proteins, the myriapods (millipedes and centipedes) strikingly have tracheal systems as well as circulating hemocyanin (Hc). In order to elucidate the evolutionary origin and biological significance of millipede Hc, we report the molecular structure (subunit composition and amino acid sequence) of multimeric (36- Supplementary material available online at

“…Tris, HEPES (Brix et al, 1994)] or determine pH from sub-samples or separately conditioned samples (e.g. Seibel et al, 1999;Weber et al, 2008) and rarely directly in the original sample (Pörtner, 1990;Zielinski et al, 2001). While added buffers prevent pH changes and may help to address specific functional characteristics of the pigment [e.g.…”

Section: Introductionmentioning

confidence: 99%

Simultaneous high-resolution pH and spectrophotometric recordings of oxygen binding in native blood microvolumes

Oellermann

Pörtner

Mark

2014

Oxygen equilibrium curves have been widely used to understand oxygen transport in numerous organisms. A major challenge has been to monitor oxygen binding characteristics and concomitant pH changes as they occur in vivo, in limited sample volumes. Here we report a technique allowing highly resolved and simultaneous monitoring of pH and blood pigment saturation in minute blood volumes. We equipped a gas diffusion chamber with a broad-range fibre-optic spectrophotometer and a micro-pH optode and recorded changes of pigment oxygenation along oxygen partial pressure (P O2 ) and pH gradients to test the setup. Oxygen binding parameters derived from measurements in only 15 μl of haemolymph from the cephalopod Octopus vulgaris showed low instrumental error (0.93%) and good agreement with published data. Broad-range spectra, each resolving 2048 data points, provided detailed insight into the complex absorbance characteristics of diverse blood types. After consideration of photobleaching and intrinsic fluorescence, pH optodes yielded accurate recordings and resolved a sigmoidal shift of 0.03 pH units in response to changing P O2 from 0 to 21 kPa. Highly resolved continuous recordings along pH gradients conformed to stepwise measurements at low rates of pH changes. In this study we showed that a diffusion chamber upgraded with a broad-range spectrophotometer and an optical pH sensor accurately characterizes oxygen binding with minimal sample consumption and manipulation. We conclude that the modified diffusion chamber is highly suitable for experimental biologists who demand high flexibility, detailed insight into oxygen binding as well as experimental and biological accuracy combined in a single setup.