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A hierarchy of disulfide-bonded subunits: the quaternary structure of Eudistylia chlorocruorin
Green, B.N.; Kuchumov, A.R.; Walz, D.A.; Moens, L.; Vinogradov, S.N. (1998). A hierarchy of disulfide-bonded subunits: the quaternary structure of Eudistylia chlorocruorin. Biochemistry (Wash.) 37(18): 6598-6605. dx.doi.org/10.1021/bi972644y
In: Biochemistry (Washington). American Chemical Society: Easton, Pa.. ISSN 0006-2960, more
Peer reviewed article  

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Keyword
    Marine

Authors  Top 
  • Green, B.N.
  • Kuchumov, A.R.
  • Walz, D.A.
  • Moens, L.
  • Vinogradov, S.N.

Abstract
    The quaternary structure of the cysteine-rich, ∼3500-kDa chlorocruorin (Chl) from the marine polychaete Eudistylia vancouverii was investigated using maximum entropy deconvolution of the electrospray ionization mass spectra (ESIMS). The native Chl provided two groups of peaks, at ∼25 and ∼33 kDa, and one peak at ∼66 kDa. ESIMS of the reduced and reduced and carbamidomethylated Chl and of its subunits obtained by HPLC provided the complete subunit composition of the Chl. Two groups of nonglobin linker chains were observed:  L1a−f (25 000.4, 25 017.9, 25 039.6, 25 057.0, 25 074.4 and 25 096.8 Da) and L2a−d (25 402.7, 25 446.0, 25 461.6 and 25 478.3Da) (±2.5 Da), with relative intensities L1:L2 = 5:2. Six globin chains were found, a1, a2, and b1−4, with reduced masses of 16 051.5, 16 172.4, 16 853.5, 17 088.9, 17 161.2 and 17 103.6 (±1.0 Da) and relative intensities of 8:4:1:4:2:1, respectively. Disulfide-bonded dimers and a tetramer of globin chains were identified:  D1 = a1 + b3 at 33 207.1; D2 at 33 374.1, which had a cysteinylated Cys (a2 + b2 + Cys); and D3 = a1 + b4 at 33 149.4 Da (±3.0 Da), with relative intensities D1:D2:D3 = 5:4:1 and T = a1 + a2 + b1 + b2 at 66 154.8 ± 4.0 Da. A 206-kDa dodecamer subunit obtained by dissociation of the Chl in 4 M urea [Qabar, A. N., et al. (1991) J. Mol. Biol. 222, 1109−1129], was found to consist only of tetramers T. A model was proposed for the Chl, based on a dimer:tetramer ratio of 2:1:  four 206-kDa dodecamers (trimer of tetramers) and 48 dimers tethered to a framework of 30 L1 and 12 L2 linker chains. The 144 globin chains (2480 kDa) and 42 linker chains (1059 kDa) provide a total mass of 3539 kDa, in good agreement with the 3480 ± 225 kDa determined previously by STEM mass mapping. The hierarchy of disulfide-bonded globin subunits observed for Eudistylia Chl provides a built-in heterogeneity of hexagonal bilayer structures.

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