生物物理化学
Online ISSN : 1349-9785
Print ISSN : 0031-9082
ISSN-L : 0031-9082
32 巻, 1 号
選択された号の論文の12件中1~12を表示しています
  • 東條 毅, 三森 経世, 岡野 裕, 小笠原 孝
    1988 年 32 巻 1 号 p. 1-6
    発行日: 1988/03/25
    公開日: 2009/03/31
    ジャーナル フリー
    By using immunoblotting technique, antigenic structures of various U small nuclear RNPs (UsnRNPs) were analysed which reacted with autoantibodies in sera from patients with collagen diseases. Based on these antigenic structures of UsnRNPs, anti-U1RNP antibodies could be more clearly differentiated from anti-U1U2U4-6RNP [Sm] antibodies. However the reactivity of anti-U1RNP antibodies with each antigenic popypeptides of U1RNP showed marked heterogeneity. An anti- (U1U2) RNP antibody system was newly described with a possible clinical association of the system.
    Immunoblotting technique was shown to be useful as a sensitive test for screening various anti-ENA antibodies in clinical laboratory. However the sensitivity of the test depended on the concentration of antigens bound to the nitrocellulose membrane.
  • 松浦 尚志, 古賀 俊逸, 宮田 康司
    1988 年 32 巻 1 号 p. 7-12
    発行日: 1988/03/25
    公開日: 2009/03/31
    ジャーナル フリー
    Using the molecular sieve high performance liquid chromatography (HPLC), we analyzed human serum apolipoproteins and attemped to isolate them. The apolipoproteins obtained from human serum very low-density lipoproteins (VLDL), high-density lipoproteins (HDL) and lipoprotein depleted serum were chromatographed on TSK G3000SW column. HPLC was performed in 6M urea (pH3.15) or 0.1M Tris-buffered 4M guanidine hydrochloride (pH7.0). As a result, better separation of apoproteins were achieved using 4M guanidine hydrochloride as an elution buffer. By this technique, apo VLDL could be separated into three peaks of apo B, apo E and apo C, apo HDL into apo E, apo A-I, apo A-II and apo C, and the apoproteins of d>1.21g/ml fraction into albumin, apo A-IV and apo A-I. Pure materials of apo E, apo A-I and apo A-IV could be obtained by a single column operation.
  • 渡部 透, 金田 惠孝, 林 泰三
    1988 年 32 巻 1 号 p. 13-16
    発行日: 1988/03/25
    公開日: 2009/03/31
    ジャーナル フリー
    It is well known that hybrids of LDH isoenzyme are formed in vitro from the mixture of LD-1 and LD-5. We have established the condition of medium to produce the hybrids, and examined the influences on the hybridization by the addition of substances concerning hypoxia. In the loading anerobic condition and in the addition of lactate and pyruvate, there were no significant effects on the hybridization and total enzyme activities. On the contrary, the addition of NADH had considerable effects upon the hybridization and enzyme activity. Namely, increasing LD-5 was markedly observed in the hybrid patterns, and the higher enzyme activity was significantly recognized for comparison with control. While, in the addition of NAD a slight increase of LD-1 was only observed in the hybrid pattern.
  • 星野 忠, 後藤 はるみ, 須藤 加代子, 内山 克己, 井川 幸雄
    1988 年 32 巻 1 号 p. 17-21
    発行日: 1988/03/25
    公開日: 2009/03/31
    ジャーナル フリー
    We have described a case of Lactate dehydrogenase (LDH) linked to IgG (kappa) immunoglobulin which LDH isoenzyme pattern shows as if six bands are present. Of these bands, one additional is located at slow gamma region and such an anomalous appearance has not previously reported. The serum contains monoclonal immunoglobulin G (kappa) of which electrophoretic mobility is at slow gamma region (same as for abnormal LDH).
    From reconstitution experiments, we concluded that all five LDH isoenzymes could bind to IgG prepared from the patient and those mobilities of the linked LDHs changed to cathodic side; LDH1, tailed to cathodic side; LDH2, moved to cathodic side; LDH3, located at slow LDH4; LDH4 located at fast LDH5; LDH5, located it slow gamma region; respectively. The affinity constant of each LDH isoenzyme was estimated and the values obtained from LDH1 to 5 were 0.026, 0.079, 0.166, 3.715, and 0.218. (Keg×109 liters/mol) was respectively. LDH4 evoked the highest affinity, it suggests that the site of antigen recognition of LDH linked to IgG was not associated with the structure of individual LDH-H and LDH-M subunits.
  • 松田 基夫, 山田 隆紹, 玉井 利孝
    1988 年 32 巻 1 号 p. 23-26
    発行日: 1988/03/25
    公開日: 2009/03/31
    ジャーナル フリー
    We obtained a new SLE-autoantibody affinity column which can fractionate U1- and U2-snRNPs simultaneously. The purified human U1- and U2-snRNPs were recovered from the column by elution with 2.5M MgCl2 at pH7.2 after the column had been washed with Tris-buffer containing 1.0M NaCl. Highly purified U1- and U2-snRNP particles were also obtained from the mouse liver (129/sv) and feline placenta, by the same procedure as used for the purification of human KB cell U1- and U2-snRNPs. By UV spectrum analysis, the materials purified from the column was suggested to form the ribonucleoprotein particles.
  • 柴田 太, 荻田 善一
    1988 年 32 巻 1 号 p. 27-31
    発行日: 1988/03/25
    公開日: 2009/03/31
    ジャーナル フリー
    我々は3種類の電気泳動法, すなわち, 等電点電気泳動法, ポリアクリルアミドゲル電気泳動法, SDSポリアクリルアミドゲル電気泳動法を組み合わせた投影二次元電気泳動法を考案し, これを用いてヒトCu, Zn型スーパーオキサイドジスムターゼ (SOD) アイソザイムを解析した。ヒトCu, Zn型SODは等電点の異なる5種のアイソザイムに分離された。これらのアイソザイムは, 非変性条件下の二次元電気泳動法により, それぞれ2~3個のスポット, 総数11個のスポットに分離された。一種類のアイソザイムから分離されるスポットのそれぞれは, 熱処理によって同一アイソザイムの他のスポットを生成することから, 異性体であると考えられる。投影二次元電気泳動法により, これらのアイソザイム, および異性体は同一の分子量(15.5kDa) を有するサブユニットから成っていることが明らかとなった。以上の結果から, これらのアイソザイムは荷電状態の差によって分離されたものであり, もとは一種類のポリペプチド鎖が後成的修飾を受けた結果生成されたものであると推定される。
  • 山本 秀子, 小島 清嗣, 真鍋 敬, 奥山 典生
    1988 年 32 巻 1 号 p. 33-38
    発行日: 1988/03/25
    公開日: 2009/03/31
    ジャーナル フリー
    For the rapid and sensitive analysis of proteins, and to obtain reproducible results, an apparatus for isotachophoresis, Shimadzu IP-2A, was modified and the electrophoretic conditions were standardized. The apparatus was modified as follows; 1) Two peristaltic pumps were used instead of N2 gas pressure system, for rinsing and refilling the capillary tube and the electrode chambers. 2) The flow lines were simplified into 3 lines, instead of 10 lines of the commercial apparatus. The electrophoretic conditions were changed as follows; 1) Hexane was overlayed on the electrolyte solutions to minimize the solubilization of CO2, instead of the use of Ba(OH)2. 2) The inner surface of the capillary tube was coated with hydroxypropyl methyl cellulose (HPMC), instead of using HPMC-containing leading electrolyte. The amount of the ampholyte mixture (Ampholine pH 3.5-10) to be applied was standardized to be 20μg. From these improvements and standardization, the interruption of electrophoresis by N2 gas bubbles or by the precipitation of BaCO3 were avoided and the procedure of isotachophoretic analysis was simplified. Serum proteins as low as 3μg could be separated into 20 UV-absorbing peaks within 16min of running time in good reproducibility.
  • 吉田 基子, 折崎 泰治, 岡野 和宣, 伊藤 迪夫
    1988 年 32 巻 1 号 p. 39-43
    発行日: 1988/03/25
    公開日: 2009/03/31
    ジャーナル フリー
    平板ゲルをガラスなどで裏打ちし補強すると, これを用いた電気泳動分離ならびに染色などの操作が容易になる. しかしながら, この裏打ちにより通常のコロイド状染料の吸着に基づく染色反応は遅くなる. 本研究ではガラス基板で裏打ちしたポリアクリルアミドゲル中に泳動分離した蛋白質にヘミンを吸着させ, そのペルオキシダーゼ様の触媒活性を利用して高速に蛋白スポットを染色する方法を開発した. すなわち, このヘミン染色法と名付けた方法では, 蛋白質に吸着したヘミンが過酸化水素の存在下で2,6-キシレノールと3,3'-ジアミノベンチジンの酸化重合反応を触媒する. この反応により蛋白質分子上で染料があらたに合成され染色される. 本研究ではこの染色反応の反応条件の最適化を行い, 45分間で蛋白質が染色されることを見出した. 本論文ではヘミン染色法とクーマシーブルー染色法の比較結果についても言及する.
  • 吉田 治弘
    1988 年 32 巻 1 号 p. 45-48
    発行日: 1988/03/25
    公開日: 2009/03/31
    ジャーナル フリー
    Capillary type isotachophoresis (CITP) and micro two-dimentional polyacrylamide gel electrophoresis (M2D-PAGE) were used by examining the genetic polymorphisms and the physiological variation of horse serum proteins. Both methods were useful for evaluating Tf typings. Especially, the CITP was a rapid analytical method. One of the very rare Tf type, Tf DM, which was undetectable by conventinal methods, was discovered by using this method. The amount of various proteins were estimated from the data on UV absorption. The Tf heterogenity was detectable by the M2D-PAGE. In addition, this method proved to be an excellent one for observing physiological conditions of each samples.
  • 黒澤 信幸, 荻田 善一
    1988 年 32 巻 1 号 p. 49-54
    発行日: 1988/03/25
    公開日: 2009/03/31
    ジャーナル フリー
    We present here a rapid method for determining substrate specificities of protease activities by using two-dimensional polyacrylamide gel electrophoresis and cellulose acetate membranes. Proteolytic active exzymes were satisfactorily separated by the two-dimensional polyacrylamide gel electrophoresis. Determining of the substrate specificities of protease activities were accomplished by using cellulose acetate membranes as absorbent of substrates. Two sheets of cellulose acetate membranes containing different substrates respectively were placed on the both sides of the thin gel layer. During incubation, enzyme activities were transferred to the membranes from the gel layer. After incubation, the membranes were stripped from the gel layer and each substrate hydrolyzing activity was detected. Comparing the zymograms, substrate specificities of proteolytic active spots were easily determined. This method is sensitive and rapid, allowing us to determine the precise substrate specificities of protease activities from only one gel layer.
  • 芝 紀代子, 戸田 年総, 長 裕子, 田中 比露美, 飯島 史朗, 大橋 望彦, 中尾 真
    1988 年 32 巻 1 号 p. 55-60
    発行日: 1988/03/25
    公開日: 2009/03/31
    ジャーナル フリー
    The apparatus, procedure and optimumized electrophoretic conditions for high-voltage isoelectric focusing on cellulose acetate membrane were reported previously. This report focuses on the steps taken to derive at the optimum conditions specified. Temperature of the membrane was maintained below 2°C even at 1500V by cooling with ice water. The membrane was kept moist by the addition of 10% (W/V) sucrose to the carrier-ampholyte solution. Pre-focusing at 500V for 30 minutes was run to obtain a pH gradient on the membrane. 30% (W/V) sucrose was added to the anode solution to prevent drying of the anode wick. The optimum conditions allowed a linear pH gradient between pH 3.5 to 8.5 to be obtained and consequently sharp protein fractionation patterns.
  • 戸沢 辰雄, 鳥居 まゆみ
    1988 年 32 巻 1 号 p. 61-63
    発行日: 1988/03/25
    公開日: 2009/03/31
    ジャーナル フリー
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