Green R L, Corotto L V, Warren G J
Advanced Genetic Sciences, Inc., Oakland, CA 94608.
Mol Gen Genet. 1988 Dec;215(1):165-72. doi: 10.1007/BF00331320.
The ice nucleation gene inaZ, from Pseudomonas syringae S203, was manipulated to produce a series of defined rearrangements in its coding sequence without changing the reading frame. The effects of these mutations on the ice nucleation phenotype were determined in a heterologous host, Escherichia coli K12. Deletions which disrupted the periodicity of 16 codons, in a repetitive region of inaZ, caused the frequencies of ice nuclei in the bacterial population to be significantly depressed; the nuclei with thresholds at warmer temperatures were most affected. In contrast, when the periodicity was left intact, deletions and duplications in the same region had only slight effects on nucleation activity. Deletions removing part or all of one of the nonrepetitive regions (that encoding the amino-terminal domain of the InaZ protein) did not abolish nucleation activity, but caused it to be limited to cooler threshold temperatures. In contrast, the non-repetitive carboxy-terminal domain of the InaZ protein was shown to be essential for ice nucleation at all temperatures. The differential requirements (for periodicity, and for the amino-terminus) in forming nuclei with different thresholds may be significant for understanding what determines the threshold temperature of an ice nucleus.
对来自丁香假单胞菌S203的冰核基因inaZ进行操作，使其编码序列产生一系列特定的重排，同时不改变阅读框。在异源宿主大肠杆菌K12中测定了这些突变对冰核表型的影响。破坏inaZ重复区域中16个密码子周期性的缺失，导致细菌群体中冰核的频率显著降低；阈值温度较高的冰核受影响最大。相反，当周期性保持完整时，同一区域的缺失和重复对成核活性只有轻微影响。去除部分或全部非重复区域（编码InaZ蛋白氨基末端结构域的区域）的缺失并没有消除成核活性，但使其仅限于较低的阈值温度。相比之下，InaZ蛋白的非重复羧基末端结构域被证明在所有温度下对冰核形成都是必不可少的。形成具有不同阈值的冰核时对周期性和氨基末端的不同要求，对于理解决定冰核阈值温度的因素可能具有重要意义。
