Hattori A, Kuroki Y, Katoh T, Takahashi H, Shen H Q, Suzuki Y, Akino T
Department of Biochemistry, Sapporo Medical University, School of Medicine, Japan.
Am J Respir Cell Mol Biol. 1996 Jun;14(6):608-19. doi: 10.1165/ajrcmb.14.6.8652189.
Pulmonary alveolar proteinosis (PAP) is a diffuse lung disease of unknown etiology in which the alveoli and terminal bronchioles of the lung fill with large amounts of surfactant-rich lipoproteinaceous materials. Its major pathologic manifestations are a small number of normal tubular myelin structures and an unusual abundance of multilamellated structures. Since surfactant protein A (SP-A) plays an important role in surfactant phospholipid homeostasis, we investigated the structural features of SP-A oligomers (alveolar proteinosis protein, APP) accumulating in the alveoli of individuals with PAP, and examined the abilities of APP to interact with lipids. Analysis of APP by Bio Gel A15m column chromatography revealed that it was composed of two protein peaks, one of which (APP-I) eluted at the position near that of blue dextran whereas the other (APP-II) eluted far behind blue dextran but ahead of thyroglobulin. These populations of APP showed almost identical amino acid compositions. Electron microscopic observations of APP molecules using the rotary shadow technique revealed that APP-II was observed as hexameric particles, presumably consisting mainly of octadecamers whose diameter was approximately 30 nm. The population seen for APP-II was similar to that seen for SP-A from healthy individuals. In contrast, APP-I was observed as multimerized larger aggregates whose diameter appeared to be about 70 to 90 nm. Both APP-I and APP-II retained the abilities to bind dipalmitoylphosphatidylcholine (DPPC). They also induced phospholipid vesicle aggregation in a concentration-dependent manner. The maximal turbidity for light scattering induced by APP-I and APP-II was almost equivalent when analyzed as a function of molar concentration. In vitro reconstitution experiments with porcine surfactant protein B (SP-B) and phospholipids revealed that the multilamellated membranes in structures formed from APP-I consisted of several layers of doubled unit membranes. APP-I failed to form tubular myelin structures. In contrast, APP-II formed well-formed lattice structures seen in tubular myelin. From these data we conclude that there exists an abnormal multimerized form of SP-A oligomer in the alveoli of patients with PAP, and that this unusual subpopulation of SP-A oligomer exhibits abnormal function on phospholipid membrane organization.
肺泡蛋白沉积症（PAP）是一种病因不明的弥漫性肺部疾病，其肺部的肺泡和终末细支气管内充满大量富含表面活性剂的脂蛋白物质。其主要病理表现为少量正常的管状髓鞘结构和异常丰富的多层结构。由于表面活性剂蛋白A（SP-A）在表面活性剂磷脂稳态中起重要作用，我们研究了在PAP患者肺泡中积累的SP-A寡聚体（肺泡蛋白沉积症蛋白，APP）的结构特征，并检测了APP与脂质相互作用的能力。通过Bio Gel A15m柱色谱分析APP，结果显示它由两个蛋白峰组成，其中一个（APP-I）在蓝色葡聚糖附近的位置洗脱，而另一个（APP-II）在蓝色葡聚糖之后但在甲状腺球蛋白之前洗脱。这些APP群体显示出几乎相同的氨基酸组成。使用旋转阴影技术对APP分子进行电子显微镜观察发现，APP-II表现为六聚体颗粒，推测主要由直径约为30nm的十八聚体组成。APP-II群体与健康个体的SP-A群体相似。相比之下，APP-I表现为多聚化的更大聚集体，其直径似乎约为70至90nm。APP-I和APP-II都保留了结合二棕榈酰磷脂酰胆碱（DPPC）的能力。它们还以浓度依赖的方式诱导磷脂囊泡聚集。当作为摩尔浓度的函数进行分析时，APP-I和APP-II诱导的光散射最大浊度几乎相等。用猪表面活性剂蛋白B（SP-B）和磷脂进行的体外重建实验表明，由APP-I形成的结构中的多层膜由几层双单位膜组成。APP-I未能形成管状髓鞘结构。相比之下，APP-II形成了在管状髓鞘中可见的结构良好的晶格结构。从这些数据我们得出结论，PAP患者的肺泡中存在SP-A寡聚体的异常多聚化形式，并且这种异常的SP-A寡聚体亚群在磷脂膜组织上表现出异常功能。
