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Phosphagen kinase of the giant tubeworm Riftia pachyptila. Cloning and expression of cytoplasmic and mitochondrial isoforms of taurocyamine kinase.

Abstract：

:The giant tubeworm Riftia pachyptila lives at deep-sea hydrothermal vents along the East Pacific Rise and the Galapagos Rift. The large size and high growth rate of R. pachyptila is supported by an endosymbiotic relationship with a chemosynthetic bacterium. Elucidation of the regulation of energy metabolism of the giant tubeworm remains an interesting problem. The purpose of this study is to determine the cDNA sequence of phosphagen kinase, one of the most important enzymes in energy metabolism, and to characterize its function. Two phosphagen kinase cDNA sequences amplified from the cDNA library of R. pachyptila showed high derived amino acid sequence identity (74%) with those of cytoplasmic taurocyamine kinase (TK) and mitochondrial TK from an annelid Arenicola brasiliensis. The cytoplasmic form of the Riftia recombinant enzyme showed stronger activity for the substrates taurocyamine and also considerable activity for lombricine (21% that of taurocyamine). The mitochondrial form, which was structurally similar to mitochondrial creatine kinase, showed stronger activity for taurocyamine, and a broader activity for various guanidine compounds: glycocyamine (35% that of taurocyamine), lombricine (31%) and arginine (3%). Both forms showed no activity for creatine. The difference in substrate specificities between the cytoplasmic and mitochondrial forms might be attributable to the large difference in the amino acid sequence of the GS region and/or several key amino acid residues for establishing guanidine substrate specificity. Based on these results, we conclude that Riftia contains at least two forms of TK as phosphagen kinase. We also report the kinetic parameters, Km and kcat, of Arenicola and Riftia TKs for the first time. The Km values for taurocyamine of Arenicola and Riftia TKs ranged from 0.9 to 4.0 mM and appear to be comparable to those of other annelid-specific enzymes, lombricine kinase and glycocyamine kinase, but are significantly lower than those of Neanthes cytoplasmic and mitochondrial creatine kinases. Comparison of kcat/Km value in various annelid phosphagen kinases indicates that Arenicola mitochondrial TK has the highest catalytic efficiency (16.2 s-1 mM-1). In Arenicola TKs, the mitochondrial form has seven-fold higher efficiency than the cytoplasmic form.

journal_name

Int J Biol Macromol

journal_title

International journal of biological macromolecules

authors

Uda K,Tanaka K,Bailly X,Zal F,Suzuki T

doi

10.1016/j.ijbiomac.2005.08.009

keywords:

subject

Has Abstract

pub_date

2005-10-30 00:00:00

pages

54-60

issue

1-2

eissn

0141-8130

issn

1879-0003

pii

S0141-8130(05)00179-0

journal_volume

pub_type

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Phosphagen kinase of the giant tubeworm Riftia pachyptila. Cloning and expression of cytoplasmic and mitochondrial isoforms of taurocyamine kinase.