Crystallization of sheep (Ovis aries) and goat (Capra hircus) haemoglobins under unbuffered low-salt conditions

Abstract

Haemoglobin is a tetrameric protein that plays a vital role in the transport of oxygen from the lungs to the tissues and of carbon dioxide back to the lungs. Even though a large amount of work has already been performed in this area, the study of the haemoglobin structures of avian and mammalian species is rather incomplete. Efforts are being made to understand the salient features of the species mentioned above. Here, whole blood plasma was collected from sheep and goat and purified by anion-exchange chromatography; the haemoglobins were crystallized by the hanging-drop vapour-diffusion method under unbuffered low-salt conditions using PEG 3350 as a precipitant. Data collection was carried out using a MAR345 image-plate detector system. Sheep haemoglobin crystallizes in the orthorhombic space group P2(1)2(1)2(1) with one whole biological molecule (alpha2beta2) in the asymmetric unit, with unit-cell parameters a = 60.231, b = 70.695, c = 131.479 A. In contrast, goat haemoglobin crystallizes in the triclinic system with two biological molecules (alpha2beta2) in the unit cell. The unit-cell parameters are a = 53.103, b = 69.382, c = 96.098 A, alpha = 110.867, beta = 91.133, gamma = 109.437 degrees.

Preliminary Crystallographic Study of Hemoglobin from Buffalo (Bubalus bubalis): A Low Oxygen Affinity Species

“…Column was eluted initially with 50mM phosphate buffer at pH 6.7, followed by 0.1M, 0.2M, 0.3M, 0.4M and 1.0M of NaCl solution with the same buffer, a single peak was obtained at 0.1M elution [8]. The homogeneity of the purified buffalo hemoglobin was checked using native-PAGE [9].…”

Structure of liganded T-state haemoglobin from cat (Felis silvestris catus), a low oxygen-affinity species, in two different crystal forms

“…The purification and crystallization of Hb has been published elsewhere (Knapp et al, 1999;Neelagandan et al, 2007;Sathya Moorthy et al, 2009;Balasubramanian et al, 2009a,b,c). Cat Hb was crystallized at room temperature in two different conditions, unbuffered low salt and buffered high salt, using the hanging-drop vapour-diffusion method.…”

Purification, Crystallization and Preliminary X-Ray Diffraction Studies on Goat (Capra hircus) Hemoglobin - A Low Oxygen Affinity Species

“…Dialyzed sample was loaded to DEAE-anion exchange chromatographic column equilibrated with 50mM of phosphate buffer at pH 6.7 [9]. Column was eluted with buffer initially, followed by stepwise increase of NaCl concentration with buffer, a single peak obtained at 0.1M elution was collected at the rate 3ml/min.…”