title: Increased yield of high purity recombinant human brain natriuretic peptide by acid hydrolysis of short fusion partner in Escherichia coli.
abstract: Recombinant human B-type natriuretic peptide (rhBNP) is a 32-amino acid peptide used to treat congestive heart failure. In this paper, we report a method for the increased production of rhBNP in Escherichia coli with high purity. hBNP was cloned with a short growth hormone fusion partner coupled with a unique acid-labile dipeptide linker to cleave the fusion protein to release the rhBNP. The recombinant fusion protein was expressed as an inclusion body (IB) and the fermentation process was optimized to produce on large scale. The IBs were recovered by cell lysis, and the pure IBs were directly treated with diluted acid to get the target peptide from the fusion protein and the resultant peptide was purified by reversed phase chromatography. The final purity of the rhBNP was more than 99% with yield of 50mg per liter of culture, which is ten times higher than the previous reports. The purified rhBNP exhibited specific biological activity similar to the standard peptide in producing cyclic-guanosine monophosphate.
authors.affiliation: Virchow Research Centre, Hyderabad 500 055, India; Centre for Biotechnology, Jawaharlal Nehru Technological University, Kukatpally, Hyderabad 500 085, India.
authors.forename: Radha Madhavi
authors.initials: RM
authors.lastname: Kanumuri
authors.affiliation: Virchow Research Centre, Hyderabad 500 055, India.
authors.forename: Chitra
authors.initials: C
authors.lastname: Bajji
authors.affiliation: Virchow Research Centre, Hyderabad 500 055, India.
authors.forename: Rajesh R
authors.initials: RR
authors.lastname: Tummuru
authors.affiliation: Department of Zoology, Yogi Vemana University, Kadapa 516 003, AP, India.
authors.forename: Venkat R R Arva
authors.initials: VR
authors.lastname: Tatireddigari
authors.affiliation: Centre for Biotechnology, Jawaharlal Nehru Technological University, Kukatpally, Hyderabad 500 085, India.
authors.forename: Lakshmi Narasu
authors.initials: LN
authors.lastname: Mangamoori
authors.affiliation: Department of Biotechnology, Govt. College for Men, Kadapa, AP, India.
authors.forename: Kalpana
authors.initials: K
authors.lastname: Panati
authors.affiliation: Department of Zoology, Yogi Vemana University, Kadapa 516 003, AP, India. Electronic address: nvramireddy@gmail.com.
authors.forename: Venkata Ramireddy
authors.initials: VR
authors.lastname: Narala
mesh_terms: D004926:Escherichia coli / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D006868:Hydrolysis; D020097:Natriuretic Peptide, Brain / Q000096:biosynthesis* / Q000737:chemistry* / Q000235:genetics / Q000302:isolation & purification*; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification
chemical_list: D011993:Recombinant Fusion Proteins; D020097:Natriuretic Peptide, Brain
keywords: Acid hydrolysis; B-type natriuretic peptide; Fusion protein; Inclusion body; Reversed phase chromatography