人血清アルブミン（遺伝子組換え）の精製に関する研究

DOI

書誌事項

タイトル別名
  • Purification of Recombinant Human Serum Albumin

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  Two major problems are expected to arise in the future in connection with plasma proteins. The first problem is the lack of source materials. The second problem is the risk of transmission of viral disease such as hepatitis, HIV or other yet-to-be identified viruses. Genetic engineering is the best approach to solve these problems. Plasma derived human serum albumin (pHSA) is one of the most useful plasma proteins. However, producing recombinant human serum albumin (rHSA) commercially via genetic engineering is not entirely straightforward. Two hurdles must be cleared in order to develop rHSA. One is cost and the other is quality. Albumin is typically administered in tens of gram quantities. At a purity level of 99.999% (a level considered sufficient for other recombinant protein preparations such as vaccine and cytokine), rHSA impurities on the order of one mg will still be injected into patients. So impurities from the host organism, in this case yeast, must be reduced to a minimum. Furthermore, purified rHSA must be identical to pHSA. Despite these stringent requirements, cost must be kept in check. One gram of pHSA costs a few dollars. Our goal is to produce rHSA at least as economically as pHSA. Thus to rein in rHSA costs, maximum quantities of albumin must be produced from minimum volumes of culture, and highly efficient, high-yield purification methods are required. Because of these special issues about rHSA development, purification methods were designed under conception described below. 1. Easy automatic control, 2. Easy scale up, 3. Use of no special materials, 4. Use of no complicated methods. The item No.1 is necessary for cost cutting and uniformity of quality. The item No. 2 is essential by the reason mentioned below. In the first stage of rHSA development, we examined purification methods based on 3 to 10 l fermentation scale. In the next stage of rHSA development, we confirmed and optimized purification methods established at the first stage using 1000 l fermentation scale. The final stage of rHSA development is commercial scale. We have the plan of fermentation scale being 50 m3. The purification methods of rHSA must be identical throughout these three stages. Easy scale up is a very important factor for process development. Column chromatography is the best approach to solve these problems. So we used a lot of column chromatography techniques in rHSA purification. The items No.3 and No.4 are necessary for the reason mentioned below. Affinity chromatography is usually a very efficient method. But in the case of rHSA, affinity chromatography is not efficient considering cost and capacity. Furthermore, gradient elution is a very efficient method for small size chromatography, but in rHSA purification, chromatography column is very large. Gel packing into large columns is much more difficult than in small ones, and also, chromatography patterns in large columns are often different from those of small ones. In other words, though impurities can be separated in small scale, separation efficiency becomes very low in large scale and thus the results of chromatography is different between large and small scales. Because of the reasons mentioned above, the media used in our process are very popular ones such as ion exchange, hydrophobic and gel filtration. The methods of chromatography we used are very simple, i.e., rHSA is adsorbed on the media, impurities are washed out, and then rHSA is eluted at a stretch. Or contrariwise, rHSA is flowed through the media while the impurities are adsorbed. In this way, scale up of chromatography is very simple. Same bed height and same linear flow give same results independent of gel volume.<br>

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詳細情報 詳細情報について

  • CRID
    1390001204473321088
  • NII論文ID
    130004851468
  • DOI
    10.11347/pda.2.28
  • ISSN
    18811728
    13444891
  • 本文言語コード
    ja
  • データソース種別
    • JaLC
    • CiNii Articles
  • 抄録ライセンスフラグ
    使用不可

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