Disease relevance of CKMT2

• Up to one third of the protein in E. coli extracts consisted of soluble recombinant Mib-CK in an enzymically active form [1].
• Sarcomeric mitochondrial creatine kinase (Mib-CK) of chicken was expressed in Escherichia coli as a soluble enzyme by using an inducible phage-T7 promoter [1].

High impact information on CKMT2

• Mib-CK consumes ATP produced in the mitochondria for the production of phosphocreatine, which is then exported into the cytosol for fast regeneration of ATP by the cytosolic CK isoforms [2].
• We have solved the structure of the octameric mitochondrial isoform, Mib-CK, which is located in the intermembrane compartment and along the cristae membranes [2].
• Finally, expression of MiaCK and MibCK was low both in situ and in vitro [3].
• B-CK and Mib-CK were localized along the entire sperm tail and in the mitochondria-rich midpiece, respectively [4].
• Free radical-induced inactivation of creatine kinase: influence on the octameric and dimeric states of the mitochondrial enzyme (Mib-CK) [5].

Biological context of CKMT2

• Recombinant Mib-CK displayed an identical N-terminal amino acid sequence, identical Km for phosphocreatine and Vmax. values, the same electrophoretic behaviour and the same immunological cross-reactivity as the native enzyme isolated from chicken heart mitochondria [1].
• The recombinant Mib-CK had the same molecular mass as native chicken Mib-CK in m.s. analysis, indicating that post-translational modification of the enzyme in chicken tissue does not occur [1].
• Selection of the Mib-CK expression plasmid on media containing kanamycin rather than ampicillin extended the time period of maximal Mib-CK expression [1].
• Replacement of Trp-206 by cysteine led to a destabilization of the active-site structure, solvent exposure of Trp-223, and to the dissociation of the Mib-CK dimers into monomers [6].

Associations of CKMT2 with chemical compounds

• The reaction of peroxynitrite (PN) with sarcomeric mitochondrial creatine kinase (Mib-CK; EC 2.7.3.2) was observed at different stages of complexity (i) with purified Mi-CK, (ii) with enzyme bound on isolated mitoplasts, and (iii) within intact respiring mitochondria [7].
• Using site-directed mutagenesis, we have substituted the corresponding reactive Cys278 in the chicken cardiac mitochondrial creatine kinase (Mib-CK) with either glycine, serine, alanine, asparagine, or aspartate [8].
• However, upon addition of MgADP plus creatine and nitrate to induce a transition-state analogue complex of the enzyme, native Mib-CK dissociated much more readily into dimers than proteinase K-digested Mib-CK [9].

Other interactions of CKMT2

• In analogy with hen egg ovalbumin, where the same proteases all cleaved the polypeptide chain very specifically around Ala-352, Ala-323 of Mib-CK may be located in an exposed surface loop that is sensitive to protease attack [9].

Analytical, diagnostic and therapeutic context of CKMT2

• Gel permeation chromatography demonstrated that the two proteolytic fragments of Mib-CK with M(r)'s of approximately 37,000 and approximately 6000 remain associated with each other [9].

References