Subunits a, b and c of Panulirus Interruptus Hemocyanin and Evolution of Arthropod Hemocyanins

Soeter, Nell M.; Beintema, Jaap J.; Jekel, Peter A.; Bak, Henk J.; Vereijken, Johan J.; Neuteboom, Ben

doi:10.1007/978-3-642-71481-8_28

Cited by 7 publications

(12 citation statements)

References 11 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…Partial or complete sequences of other parts have been published as well, but without the evidence [4, 6,8,13, 141. In this paper, the complete evidence for the amino acid sequence of the 55-kDa fragment will be given, completing the primary-structure determination of subunit a.…”

mentioning

confidence: 99%

Panulirus interruptus hemocyanin. The elucidation of the complete amino acid sequence of subunit a

Bak

Beintema

1987

Eur J Biochem

Self Cite

View full text Add to dashboard Cite

As a final step in the elucidation of the primary structure of subunit a of Panulirus interruptus hemocyanin (657 residues, M , 75696 excluding two copper ions and carbohydrate), the amino acid sequence of the largest fragment obtained by limited trypsinolysis was determined. The elucidation of the sequence of residues 176 -657, comprising domains two and three, was mainly based on two digests, with CNBr and trypsin, respectively, from both of which a complete set of peptides was obtained. Additional sequence information was obtained from a digest with Staphylococcus aureus V8 protease and from one fragment obtained by cleaving subunit a with hydroxylamine. A block during Edman degradations indicated an Asn-Gly sequence at positions 597 -598, although only aspartic acid was identified at position 597.Hemocyanins are large multi-subunit copper-containing oxygen carriers occurring freely dissolved in the hemolymph of many molluscs and arthropods. Arthropod hemocyanins are composed of hexamers or multihexamers with subunits of about 75000 daltons, each of which contains one binuclear copper site [l, 21. Like in all arthropods [l, 21, hemocyanin of the spiny lobster Panulirus interruptus displays subunit heterogeneity [3]. There are at least three subunit types, named a, b and c, which were formerly called '94K', '90K' and '80K', respectively, since their apparent molecular masses on SDS/ polyacrylamide gels were 94, 90 and 80 kDa [3, 41. These subunits have been characterized by examination of their electrophoretic, chromatographic and immunological properties [3, 51. Subunits a and b, accounting for 80-9o0/0 of the monomers upon dissociation of the hexamers, are very similar, but subunit c is quite different [3, 6 -81. Both X-ray diffraction studies and amino acid sequence studies are being performed. The three-dimensional structure has been determined at a resolution of 0.32 nm with crystals containing a mixture of subunits a and b in roughly equal amounts [4, 93. Each polypeptide chain is folded into three distinct domains.Primary-structure investigations were started with subunit a, which gave rise to three fragments upon limited trypsinolysis : an N-terminal 18-kDa fragment, a C-terminal 55-kDa fragment (formerly called 71-kDa fragment after its apparent molecular mass on SDS-PAGE) and a glycopeptide located in between [lo]. The 18-kDa fragment and the glycopeptide together form the first domain while the 55-kDa fragment accounts for domains two and three. The second domain contains the active site, with two copper atoms. Each copper atom is ligated by three histidines, two of which are provided by a -His-Xaa-Xaa-Xaa-His-sequence located in a helix, the third coming from another helix.The sequences of the first 230 residues, comprising the 18-kDa fragment, the glycopeptide, and of 55 N-terminal residues of the 55-kDa fragment have already been reported [lo-121. Partial or complete sequences of other parts have been published as well, but without the evidence [4, 6,8,13, 141. In this paper, the complete eviden...

show abstract

mentioning

confidence: 99%

Panulirus interruptus hemocyanin. The elucidation of the complete amino acid sequence of subunit a

Bak

Beintema

1987

Eur J Biochem

Self Cite

View full text Add to dashboard Cite

show abstract

“…Isolation procedures and amino acid analyses of the cyanogen bromide and tryptic peptides are presented in Tables 1 and 2. Additional experimental evidence (gel filtration patterns, isolation procedures and amino acid compositions of other peptides generated by secondary digestion of the tryptic peptides) are given in [21]. Analytical data of these latter peptides are com- Fig.…”

Section: Results and Discussion The Primary Structure O F The 18-kda mentioning

confidence: 99%

“…The N-terminal sequences of the three subunits have been published [4]. Subunits a and b are very similar, subunit c is quite different [5].Limited tryptic cleavage of subunit a gives rise to three fragments: an N-terminal 18-kDa fragment, a C-terminal 55-kDa fragment and a glycopeptide of 13 amino acid residues located in between [6].Both the primary and the three-dimensional structures are under investigation. The three-dimensional structure has been determined at a resolution of 0.32 nm.…”

mentioning

confidence: 99%

See 1 more Smart Citation

Primary and tertiary structures of the first domain of Panulirus interruptus hemocyanin and comparison of arthropod hemocyanins

Soeter¹,

Jekel²,

Beintema³

et al. 1987

Eur J Biochem

Self Cite

View full text Add to dashboard Cite

The amino acid sequence of the first domain (positions 1 -175) of Panulirus interruptus hemocyanin subunit a has been determined. The sequence of residues 1 -158 (18-kDa fragment obtained by limited proteolysis) was derived from peptides obtained by digestion of this fragment with CNBr and trypsin and by subdigestion of these peptides with other enzymes. The peptides were sequenced automatically or manually. The amino acid sequence has been fitted into the electron-density map at 0.32-nm resolution.The residues of domain 1 are folded into a larger, mainly helical, globular part, containing one disulfide bridge, and a smaller part near the molecular twofold axis. The latter part consists of an c( helix and a fl strand which contains a covalently attached carbohydrate moiety. The sites susceptible to limited proteolytic cleavage of the subunit are discussed.Comparison of the N-terminal sequence with those of other arthropod hemocyanins revealed, besides an Nterminal extension of five residues, the presence of a 21-residue loop (positions 22 -42) in the crustacean sequences. This loop contains helix 1.2, a less defined region in the electron-density map. It is absent in chelicerate sequences. Strong evidence is presented that: (a) the structure of the first 21 residues (including helix 1.1) is the same in all arthropod hemocyanins with known amino acid sequence; (b) a stretch containing about 15 residues (including part of helix 1.3) following the 21-residue loop has a different structure in crustaceans and chelicerates; (c) the rest of domain 1 has the same structure again. It is shown that all conserved residues are in the contact region with the other two domains.Hemocyanins are copper-containing oxygen carriers occurring in the hemolymph of arthropods and molluscs. Arthropod hemocyanins consist of hexamers or multihexamers [I, 21. In the spiny lobster, Panulirus interruptus, hemocyanin molecules are present as hexamers built from three different subunits, called a, b and c, each with a molecular mass of about 75 kDa [3]. These were formerly called 94 K, 90 K and 80 K, respectively, according to their apparent molecular mass on SDS/polyacrylamide gels. The N-terminal sequences of the three subunits have been published [4]. Subunits a and b are very similar, subunit c is quite different [5].Limited tryptic cleavage of subunit a gives rise to three fragments: an N-terminal 18-kDa fragment, a C-terminal 55-kDa fragment and a glycopeptide of 13 amino acid residues located in between [6].Both the primary and the three-dimensional structures are under investigation. The three-dimensional structure has been determined at a resolution of 0.32 nm. It is the only structure of an arthropod hemocyanin in which the polypeptide chain has been traced. The polypeptide chain, which is 657 residues Correspondence to N. M . Soeter, Rudolf Magnus Instituut voor

show abstract

“…Also a plastic regulation of cooperative transition is provided by subunit heterogeneity as shown in P. interruptus. This Hc is naturally composed by a mixture of three distinct subunits, namely a, b and c, (Neuteboom et al, 1992), but the last does not participate in the cooperativity of the hexamers (Soeter et al, 1986). Homohexamers prepared in vitro from isolated subunits have in general different affinity or cooperativity as compared with the native heterohexamers (Dainese et al, 1998;Molon et al, 2000).…”

Section: Subunit Diversity and Oxygen-binding Modulationmentioning

confidence: 99%

The molecular heterogeneity of hemocyanin: Its role in the adaptive plasticity of Crustacea

Giomi

Beltramini

2007

Gene

View full text Add to dashboard Cite

Subunits a, b and c of Panulirus Interruptus Hemocyanin and Evolution of Arthropod Hemocyanins

Cited by 7 publications

References 11 publications

Panulirus interruptus hemocyanin. The elucidation of the complete amino acid sequence of subunit a

Panulirus interruptus hemocyanin. The elucidation of the complete amino acid sequence of subunit a

Primary and tertiary structures of the first domain of Panulirus interruptus hemocyanin and comparison of arthropod hemocyanins

The molecular heterogeneity of hemocyanin: Its role in the adaptive plasticity of Crustacea

Contact Info

Product

Resources

About