New regulation mechanism of polyamine biosynthesis mediated by ribosomal Protein, L10 as an antizyme

瘤胃单胞菌是一种严格厌氧的革兰氏阴性细菌，它既没有游离型的脂蛋白，也没有结合型的脂蛋白。在一般的革兰氏阴性菌中，脂蛋白在维持细胞表面的结构完整性方面起着重要的作用。相反，它含有身体碱，它与肽聚糖共价连接，作为细胞分裂的关键成分。在S.rmuinantimn中，身体碱是由L赖氨酸在赖氨酸脱羧酶(LDC)的作用下合成的，并作为正常细胞生长所必需的成分转移到肽聚糖中。此外，苏云金杆菌还具有两个独特的特性：(1)对L赖氨酸和L鸟氨酸均有脱羧酶活性。(2)生物化学前景与真核鸟氨酸相似。在这里，我们发现了以下证据：(1)LDC中的S.RTM在静止的早期被三磷酸腺苷依赖的丝氨酸蛋白酶(S)显著降解。这种降解蛋白质的降解是必需的降解事实…更多或更多，被命名为P22。P22是由腐胺在兰氏链霉菌细胞中积累诱导的，它与LDC紧密结合，K_D=8.5×10~(-11)&gt；M。P22似乎具有与抗Zynte(AZ)相似的生化和生物物理特性，这是仅在哺乳动物细胞中报道的。P22为Rimliulimn核糖体蛋白Li0。L10与LDC结合形成LDC-L10复合体可能引起LDC亚单位的构象改变(S)，使其被三磷酸腺苷依赖的丝氨酸蛋白酶(S)攻击，使LDC被分解成小肽。我们还鉴定了L-10分子中与LDC结合所必需的两个区域，A区(K^&lt；101&gt；NKLD^&lt；105&gt；)和B区(G^&lt；106&gt；VIRNAVYVLD^&lt；170&gt；)。(2)我们鉴定了参与反刍链霉菌LDC降解的依赖三磷酸腺苷的丝氨酸蛋白酶(S)。瘤胃沙门氏菌中存在三个依赖三磷酸腺苷的蛋白水解酶基因，其中一个属于C IpXP(S)，因为它们需要一个适配蛋白。CIpXP蛋白酶由CIpX ATPase和CIPP多肽酶组成。在S.rmineantitan中，L10可能作为一种适配蛋白，携带LDC到CIpXP酶。我们将反刍链霉菌C1pP基因克隆到pET15b中，并在大肠杆菌BL21(DE3)中表达和纯化。(3)主要外膜蛋白Mep45的肽聚糖与S层同源结构域之间的相互作用需要与肽聚糖共价连接的身体碱。在这里，使用一系列一般结构为NH_3^(CH_2)_nNH_3^，n=3-6的二胺，我们发现身体碱(n=5)是获得细胞对外部损伤剂的高抵抗力的肽聚糖中最有效的成分，并且是Mep45的SLH结构域与肽聚糖之间有效相互作用所必需的，有助于外膜正确地锚定到肽聚糖上。较少

Selenomonas ruminantium, which is a strictly anaerobic, Gram-negative bacterium has neither free nor bound forms of lipoprotein which plays an important role of the maintenance of the structural integrity of the cell surface in general Gram-negative bacteria. Instead, it has cadaverine, which links covalently to the pepticloglycan as a pivotal constituent for the cell devision. In S. rmuinantimn cadaverine is synthesized constitutively from L-lysine by lysine decarboxylase (LDC) and transferred to the peptidoglycan as an essential constituent for normal cell growth. Furthermore, S. rwninantuan LDC has two unique characteristics ; (1) it has decarboxylase activity towards both L-lysine and L-ornithine. (2) biochemical future is similar to that of eukaryotic ornithine. Here, we found the following evidences, i.e.(1) S. rtm inanluon LDC was dramatically degraded at the early stationary phase by ATP -dependent serine protease(s). This degradation proteolysis was required a degradation fact … More or, which was named as P22. P22 was induced by the accumulation of putrescine in S. raminanlium cells, and it bound tightly to LDC with a K_D) of 8.5X10^<-11> M. P22 seems to have similar biochemical and biophysical characteristics to those of an antizynte (AZ), which have been reported only in mammalian cells. P22 was identified as a ribosomal protein LI0 of S. rimliuunlimn. The binding of L10 to LDC for forming LDC-L10 complex may cause a conformational change(s) of LDC subunit, which allow it to be attacked by ATP-dependent serine protease(s), and the LDC would be broken down to small peptides. We also identified the two regions, A region (K^<101>NKLD^<105>) and B region (G^<106>VIRNAVYVLD^<170>) in L-10 molecule essential for its binding to LDC.(2) We identified the ATP-dependent serine protease(s), which is involved in the degradation of S. ruminantiian LDC. There were three genes for ATP-dependent protease chromosomal in S. ruminantium and we focused on one of them, which belongs to C IpXP protease(s) since they require an adapter protein. CIpXP protease consists of the CIpX ATPase and the CIpP peptidase. In S. rmninantitan, L10 may work as an adapter protein and carry the LDC to CIpXP protease. We cloned C1pP gene from S. ruminantiuni into pET15b and expressed it in E, coli BL21 (DE3) and purified.(3) Cadaverine covalently linked to the peptidoglycan is required for the interaction between the peptidoglycan and the S-layer homologous (SLH) domain of the major outer membrane protein, Mep45. Here, using a series of diamines with a general structure of NH_3^+(CH_2)_nNH_3^+, n=3-6), we found that cadaverine (n=5) specifically serves as the most efficient constituent of the peptidoglycan to acquire the high resistance of the cell to the external damage agents, and is required for the effective interaction between the SLH domain of Mep45 and the peptidoglycan facilitating the correct anchoring of the outer membrane to the peptidoglycan. Less