A rapid and cost-effective method of producing recombinant proBNP and NT-proBNP variants in Escherichia coli for immunoassay of heart failure

作者：Soleh Muhammad Tarmizi; Foo Jared Yong Yang; Bailey Ulla Maja; Tan Nikki Yi; Wan Yunxia; Cooper White Justin; Schulz Benjamin Luke; Punyadeera Chamindie^*

来源：Biotechnology Letters, 2014, 36(1): 133-140.

DOI：10.1007/s10529-013-1341-0

• AlphaLISA
• Escherichia coli
• Heart failure
• Immunoassay
• Natriuretic peptides
• Plasma peptides

The measurements of plasma natriuretic peptides (NT-proBNP, proBNP and BNP) are used to diagnose heart failure but these are expensive to produce. We describe a rapid, cheap and facile production of proteins for immunoassays of heart failure. DNA encoding N-terminally His-tagged NT-proBNP and proBNP were cloned into the pJexpress404 vector. ProBNP and NT-proBNP peptides were expressed in Escherichia coli, purified and refolded in vitro. The analytical performance of these peptides were comparable with commercial analytes (NT-proBNP EC50 for the recombinant is 2.6 ng/ml and for the commercial material is 5.3 ng/ml) and the EC50 for recombinant and commercial proBNP, are 3.6 and 5.7 ng/ml respectively). Total yield of purified refolded NT-proBNP peptide was 1.75 mg/l and proBNP was 0.088 mg/l. This approach may also be useful in expressing other protein analytes for immunoassay applications.
To develop a cost effective protein expression method in E. coli to obtain high yields of NT-proBNP (1.75 mg/l) and proBNP (0.088 mg/l) peptides for immunoassay use.

• 出版日期2014-1