The S-layer protein of Lactobacillus acidophilus ATCC 4356:: Identification and characterisation of domains responsible for S-protein assembly and cell wall binding
The S-layer protein of Lactobacillus acidophilus ATCC 4356:: Identification and characterisation of domains responsible for S-protein assembly and cell wall binding
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DOI：
10.1006/jmbi.2000.4258
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发表时间：
2001-01-12
影响因子：
5.6
通讯作者：
Pouwels, PH
中科院分区：
文献类型：
--
作者：
Smit, E;Oling, F;Pouwels, PH
嗜酸乳杆菌与许多其他细菌一样，具有由43 kDa的蛋白质（S-A-蛋白）组成的表面层。S-A-蛋白可以很容易地提取和结晶成大的结晶补丁在脂质单层上的净负电荷，但不是在脂质上的净中性电荷。从在二油酰磷脂酰丝氨酸上生长的晶体重建S-层表明具有单位晶胞尺寸（a = 118埃; B = 53埃，和γ = 102度）的倾斜晶格，类似于瑞士乳杆菌ATCC 12046的S-层所确定的那些。S-A蛋白与L.瑞士乳杆菌（Lactobacillus helveticus）、卷曲乳杆菌（Lactobacillus crispatus）的S蛋白基因编码的S蛋白。嗜酸乳杆菌和卷曲乳杆菌提出存在两个结构域，一个结构域包含S-A-蛋白的N-末端三分之二（SAN），另一个结构域由S-A-蛋白的C-末端三分之一（SAC）组成。N-末端结构域的序列是可变的，而C-末端结构域的序列在这些生物体的S-蛋白中是高度保守的，并且包含串联重复。对S-A-蛋白的酶切分析表明，SAN具有蛋白酶抗性，结构紧凑。SAC被蛋白酶迅速降解，因此可能具有更易接近的结构。编码SAN或绿色荧光蛋白与SAC融合（GFP-SAC）的DNA序列在大肠杆菌中有效表达。纯化的SAN可以结晶成单层和多层晶体，具有与真实S-A蛋白相同的晶格参数。计算的S-A-蛋白减去SAN密度差图揭示了SAC结构域在投影中的可能位置，该SAC结构域在截短的SAN肽中缺失。GFP-SAC融合蛋白与L.嗜酸乳杆菌L. helveticus和L.卷曲细胞的S-层已被删除，但不是非剥离的细胞或干酪乳杆菌。(C)北京：科学出版社.
Lactobacillus acidophilus, like many other bacteria, harbors a surface layer consisting of a protein (S-A-protein) of 43 kDa. S-A-protein could be readily extracted and crystallized in vitro into large crystalline patches on lipid monolayers with a net negative charge but not on lipids with a net neutral charge. Reconstruction of the S-layer from crystals grown on dioleoylphosphatidylserine indicated an oblique lattice with unit cell dimensions (a = 118 Angstrom; b = 53 Angstrom, and gamma = 102 degrees) resembling those determined for the S-layer of Lactobacillus helveticus ATCC 12046. Sequence comparison of S-A-protein with S-proteins from L. helveticus, Lactobacillus crispatus and the S-proteins encoded by the silent S-protein genes from L. acidophilus and L, crispatus suggested the presence of two domains, one comprising the N-terminal two-thirds (SAN), and another made up of the C-terminal one-third (SAC) of S-A-protein. The sequence of the N-terminal domains is variable, while that of the C-terminal domain is highly conserved in the S-proteins of these organisms and contains a tandem repeat. Proteolytic digestion of S-A-protein showed that SAN was protease-resistant, suggesting a compact structure. SAC was rapidly degraded by proteases and therefore probably has a more accessible structure. DNA sequences encoding SAN or Green Fluorescent Protein fused to SAC (GFP-SAC) were efficiently expressed in Escherichia coli. Purified SAN could crystallize into mono and multi-layered crystals with the same lattice parameters as those found for authentic S-A-protein. A calculated S-A-protein minus SAN density-difference map revealed the probable location, in projection, of the SAC domain, which is missing from the truncated SAN peptide. The GFP-SAC fusion product was shown to bind to the surface of L. acidophilus, L. helveticus and L. crispatus cells from which the S-layer had been removed, but not to non-stripped cells or to Lactobacillus casei. (C) 2001 Academic Press.