Crystallization and Preliminary Analysis of Crystals of the 24-Meric Hemocyanin of the Emperor Scorpion (Pandinus imperator)

Abstract

Hemocyanins are giant oxygen transport proteins found in the hemolymph of several invertebrate phyla. They constitute giant multimeric molecules whose size range up to that of cell organelles such as ribosomes or even small viruses. Oxygen is reversibly bound by hemocyanins at binuclear copper centers. Subunit interactions within the multisubunit hemocyanin complex lead to diverse allosteric effects such as the highest cooperativity for oxygen binding found in nature. Crystal structures of a native hemocyanin oligomer larger than a hexameric substructure have not been published until now. We report for the first time growth and preliminary analysis of crystals of the 24-meric hemocyanin (MW = 1.8 MDa) of emperor scorpion (Pandinus imperator), which diffract to a resolution of 6.5 Å. The crystals are monoclinc with space group C 1 2 1 and cell dimensions a = 311.61 Å, b = 246.58 Å and c = 251.10 Å (α = 90.00°, β = 90.02°, γ = 90.00°). The asymmetric unit contains one molecule of the 24-meric hemocyanin and the solvent content of the crystals is 56%. A preliminary analysis of the hemocyanin structure reveals that emperor scorpion hemocyanin crystallizes in the same oxygenated conformation, which is also present in solution as previously shown by cryo-EM reconstruction and small angle x-ray scattering experiments.

Identifying candidate genes involved in the regulation of growth using comparative transcriptome analysis (Androctonus crassicauda)

“…Recently studies suggested that Tachylectins plays an important role as pattern recognition molecules in the innate immune response system and blood coagulation ((Gokudan et al, 1999). Homocyanins, a large copper-binding metalloproteins in the hemolymph of invertebrates, acts as a respiratory pigment in the transport of oxygen and responsible for the blue coloration of arthropods hemolymph (Hosseinzadeh et al, 2019;Jaenicke et al, 2012). The present studies showed that, even though the levels of Hemocyanin mRNA were found in all samples but the highest level of Hemocyanin mRNA expression occurred in the adult scorpions.…”

“…region or in the visible range of the spectrum (Padmanabhanaidu, 1966). Although, the preliminary studies on the physiological properties and structural of the Haemocyanin in scorpions has been con rmed the respiratory and also buffering function of Haemocyanin (Padmanabhanaidu, 1966;Jaenicke et al, 2012), none of them has been compared the expression of Homocyanin in adult and juvenile scorpions and this issue needs further investigation.…”

Inter-Subunit Dynamics Controls Tunnel Formation During the Oxygenation Process in Hemocyanin Hexamers

“…Hemocyanins (HCNs) are large type-3 metalloproteins that transport oxygen in the blood of invertebrates like arthropods and molluscs ( Van Holde and Miller, 1995 ; Terwilliger, 1998 ; Burmester, 2002 ; Pick et al, 2008 ; Rehm et al, 2012 ; Marxen et al, 2014 ; Pinnow et al, 2016 ). Their binuclear active site coordinates copper as metal and accommodates oxygen as a peroxide ion, due to which copper oxidizes from (I) to (II) oxidation state ( Himmelwright et al, 1978 , 1980 ; Magnus and Ton-That, 1991 ; Jaenicke et al, 2012 ). Besides oxygen transportation, they have also been found to play a major role in diverse physiological functions like homeostasis, transportation of hormones ( Prosser and Brown, 1961 ), in the development of immunity via activation of phenoloxidases protein that plays a major role in the formation of their exoskeleton formation and melamine pigments, as well as wound recovery ( Lei et al, 2008 ; Coates et al, 2011 ; Coates and Nairn, 2014 ; Zhong et al, 2016 ; Coates and Talbot, 2018 ).…”

“…Therefore, a method that could provide microscopic insights into the dynamic behavior of the HCNs and variations as a function of time needs to be employed. Many experimental studies have been reported for HCNs (Decker et al, 1988;Volbeda and Hol, 1989;Magnus and Ton-That, 1991;Hazes et al, 1993;Magnus et al, 1994;Jaenicke et al, 2012;Saito and Thiel, 2014;Naresh et al, 2015Naresh et al, , 2015Bux et al, 2018). Most of the up-to-date work commonly discusses the structural features of protein in its monomeric form and rarely in its biologically active hexameric form (Volbeda and Hol, 1989).…”

Myriapod haemocyanin: the first three-dimensional reconstruction ofScolopendra subspinipesand preliminary structural analysis ofS. viridicornis

“…70-75 kDa) consists of three domains: domain I contains five to six α-helices, domain II contains a four α-helix bundle and domain III is a seven stranded anti-parallel β-barrel. Domain II also encompasses the di-copper centre [14,[28][29][30][31][32]. The geometry and coordination environment of the active site of the arthropod PO are very similar to those of the arthropod Hc; and while not all POs are hexameric [33], those from crustaceans are [34,35].…”