Conformational transmission in proinsulin and its derivatives: a study using H/D exchange
Younas, Hooria, Gardner, Qurra-tul-Ann Afza, Rashid, Naeem, Wright, J. Neville and Akhtar, Muhammad (2011) Conformational transmission in proinsulin and its derivatives: a study using H/D exchange. International Journal of Mass Spectrometry, 302, (1-3), 36-43. (doi:10.1016/j.ijms.2010.07.020).
Buffalo proinsulin cDNA was isolated, sequenced and shown to differ from that of its bovine counterpart in six nucleotides. However, at the protein level the predicted sequences of the two species are identical. Buffalo M-proinsulin, containing the initiation methionine, was produced in Escherichia coli and purified to give Mr of 8812. Following the replacement of 99% of the exchangeable hydrogen atoms with deuterons a preparation containing 131 D atoms was obtained. Buffer exchange of the latter into a protio medium led to, the immediate release of 109 (±1) D atoms into the medium and the retention of 22 (±1) D atoms in the protein. The slow exchange of these D atoms was studied at 0 °C/pH 2.8. Insulin derived from buffalo proinsulin as well as bovine when deuteriated and buffer exchanged, similarly, gave the retention of 25 (±1) D atoms. The data show that the secondary structure of the insulin core present within buffalo/bovine proinsulin contains 5 (±1) fewer slow exchanging hydrogen atoms than are present in the final hormone. This effect is attributed, predominantly, to the long range influence of the C-peptide, composed of 26 residues, on the insulin core of buffalo proinsulin. In contrast, in the case of human proinsulin, comprising 31 amino acids in the C-peptide, the secondary structure of the insulin core within human proinsulin is closer to that of insulin itself.
|Keywords:||buffalo/bovine proinsulin, expression, refolding and mass spectrometry, proinsulin and insulin replacement of exchangeable hydrogen atoms with deuterons, proinsulin–insulin structure comparison, long range and proximal effects in buffalo proinsulin and derivatives|
|Subjects:||Q Science > Q Science (General)
Q Science > QD Chemistry
Q Science > QH Natural history > QH301 Biology
|Divisions:||Faculty of Natural and Environmental Sciences > Biological Sciences
|Date Deposited:||13 Mar 2012 12:37|
|Last Modified:||27 Mar 2014 20:19|
|RDF:||RDF+N-Triples, RDF+N3, RDF+XML, Browse.|
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