TY - JOUR
T1 - Recovery and primary purification of bacteriophage M13 using aqueous two-phase systems
AU - González-Mora, Alejandro
AU - Ruiz-Ruiz, Federico
AU - Benavides, Jorge
AU - Willson, Richard C.
AU - Rito-Palomares, Marco
N1 - Funding Information:
The authors acknowledge the financial support of Tecnológico de Monterrey, Bioprocess and Synthetic Biology Research Chair (Grant CAT161) and to CONACyT for the fellowship of Alejandro González Mora (No.708102). This work was funded in part by grants by the NIAID/NIH (1R21AI111120-01A1) and the NSF (CBET-1511789).
Publisher Copyright:
© 2017 Society of Chemical Industry
PY - 2017/11
Y1 - 2017/11
N2 - BACKGROUND: Bacteriophage M13 is an Escherichia coli-specific non-lytic filamentous virus commonly used in applications ranging from antibody screening and nanomaterial construction to drug delivery, among others. In this tenor, alternative methods for the fractionation, recovery and partial purification of phage particles are desired. In this work, the use of aqueous two-phase systems (ATPS) was evaluated as an alternative method for the recovery of phage particles. RESULTS: The partition behavior of M13 in PEG–salt and ionic liquid (IL)–salt ATPS was characterized using a pre-purified feedstock. In PEG-salt ATPS, M13 was preferentially partitioned to the interface. In IL ATPS, however, M13 showed a high-top phase preference with recovery yields above 65%. Selected systems were tested for the extraction of M13 from a crude fermentation broth. From crude broth, a PEG 400-potassium phosphate system with volume ratio (VR) of 1 and 25% w/w tie line length (TLL) gave the best M13 top phase recovery (83%) and purification fold (18.2) in terms of total protein concentration. CONCLUSIONS: The results presented here demonstrate the practical application of ATPS as an efficient process for the primary recovery and partial purification of M13 and represent the first study of the extraction of viral particles directly from a crude broth as well as the use of IL-Salt ATPS.
AB - BACKGROUND: Bacteriophage M13 is an Escherichia coli-specific non-lytic filamentous virus commonly used in applications ranging from antibody screening and nanomaterial construction to drug delivery, among others. In this tenor, alternative methods for the fractionation, recovery and partial purification of phage particles are desired. In this work, the use of aqueous two-phase systems (ATPS) was evaluated as an alternative method for the recovery of phage particles. RESULTS: The partition behavior of M13 in PEG–salt and ionic liquid (IL)–salt ATPS was characterized using a pre-purified feedstock. In PEG-salt ATPS, M13 was preferentially partitioned to the interface. In IL ATPS, however, M13 showed a high-top phase preference with recovery yields above 65%. Selected systems were tested for the extraction of M13 from a crude fermentation broth. From crude broth, a PEG 400-potassium phosphate system with volume ratio (VR) of 1 and 25% w/w tie line length (TLL) gave the best M13 top phase recovery (83%) and purification fold (18.2) in terms of total protein concentration. CONCLUSIONS: The results presented here demonstrate the practical application of ATPS as an efficient process for the primary recovery and partial purification of M13 and represent the first study of the extraction of viral particles directly from a crude broth as well as the use of IL-Salt ATPS.
KW - aqueous two-phase systems
KW - bacteriophage M13
KW - ionic liquid
KW - partitioning
KW - plaque forming assay, purification
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U2 - 10.1002/jctb.5359
DO - 10.1002/jctb.5359
M3 - Article
AN - SCOPUS:85026729050
VL - 92
SP - 2808
EP - 2816
JO - Journal of Chemical Technology and Biotechnology
JF - Journal of Chemical Technology and Biotechnology
SN - 0268-2575
IS - 11
ER -