The objective of this study was to establish a one-step chromatographic process, by optimizing the chromatographic conditions, for purifying recombinant human type Ⅲ collagen from Pichia pastoris fermentation broth. Collagen was precipitated from fermentation broth using 30% (mass fraction) saturated ammonium sulfate. Then, the medium was ultrafiltered to 20 mmol/L phosphate buffer (PB) (pH 6.0) after reconstitution of the precipitate with ultrapure water. Following this, a SP Sepharose HP cation exchange column was used for purification, and heteroproteins were washed with 20 mmol/L PB (pH 6.0)-150 mmol/L NaCl. Finally, recombinant human type Ⅲ collagen was eluted with 20 mmol/L PB (pH 6.0)-250 mmol/L NaCl. Using this process to purify recombinant human type Ⅲ collagen from 60 L fermentation broth, the harvested product had a purity of 97.7% and total recovery rate of 68.8%. The purified recombinant human type Ⅲ collagen exhibited high purity and low pyrogen content and thus can be used in medicine industry. This study established a one-step chromatography purification process for the efficient preparation of recombinant human type Ⅲ collagen from Pichia pastoris fermentation broth. The process is simple, stable, and reliable. Ion exchange chromatography is easy to scale up, laying a foundation for industrial production of recombinant human type Ⅲ collagen.
LIANG Xin
,
ZHANG Renhuai
,
LYU Zili
,
AI Huawei
,
LIU Dong
,
LIANG Bo
,
SHAN Xudong
,
CHEN Haoran
. Isolation and purification of recombinant human type Ⅲ collagen[J]. Food and Fermentation Industries, 2020
, 46(16)
: 159
-163
.
DOI: 10.13995/j.cnki.11-1802/ts.023967
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