Fett J W, Strydom D J, Lobb R R, Alderman E M, Vallee B L, Artymiuk P J, Collett S, Phillips D C, Dobson C M, Redfield C
Biochemistry. 1985 Feb 12;24(4):965-75. doi: 10.1021/bi00325a024.
One of the major proteins secreted by an established human colon adenocarcinoma cell line has been isolated in 25% yield from the serum-free medium in which the cells were grown and identified as lysozyme. Its purification was achieved by sequential steps of acidification, cation-exchange chromatography, and reversed-phase high-performance liquid chromatography. It was recognized to be a human lysozyme on the basis of its molecular weight (14 000), isoelectric point (10.5), amino acid composition, and enzymatic activity. Its identity with previously characterized human lysozymes was established by amino-terminal sequence, peptide composition, immunological properties, NMR, and crystallography. A 4-day, 7-L collection of conditioned medium contained 20.3 mg of secreted protein of which 4.9 mg or approximately 24% of the total was tumor-derived lysozyme. The intracellular level of lysozyme was approximately 18 ng per 10(6) carcinoma cells. The possible significance of these findings in regard to the malignant process and tumor maintenance is discussed.
从一种已建立的人结肠腺癌细胞系分泌的主要蛋白质之一,已从该细胞生长的无血清培养基中以25%的产率分离出来,并被鉴定为溶菌酶。通过酸化、阳离子交换色谱和反相高效液相色谱等连续步骤实现了其纯化。基于其分子量(14000)、等电点(10.5)、氨基酸组成和酶活性,它被确认为人溶菌酶。通过氨基末端序列、肽组成、免疫特性、核磁共振和晶体学确定了它与先前表征的人溶菌酶的一致性。4天收集的7升条件培养基含有20.3毫克分泌蛋白,其中4.9毫克或约占总量24%的是肿瘤来源的溶菌酶。每10⁶个癌细胞中溶菌酶的细胞内水平约为18纳克。讨论了这些发现对于恶性过程和肿瘤维持的可能意义。