Hydrophobin-1 promotes thermostability of firefly luciferase
نویسندگان
چکیده
منابع مشابه
SpyTag/SpyCatcher Cyclization Enhances the Thermostability of Firefly Luciferase
SpyTag can spontaneously form a covalent isopeptide bond with its protein partner SpyCatcher. Firefly luciferase from Photinus pyralis was cyclized in vivo by fusing SpyCatcher at the N terminus and SpyTag at the C terminus. Circular LUC was more thermostable and alkali-tolerant than the wild type, without compromising the specific activity. Structural analysis indicated that the cyclized LUC i...
متن کاملImproved thermostability of the North American firefly luciferase: saturation mutagenesis at position 354.
We have used random chemical mutagenesis and a simple genetic screen to generate and isolate a thermostable mutant of luciferase from the North American firefly (Photinus pyralis). A single G-to-A transition mutation, resulting in the substitution of a glutamate for a lysine residue at position 354 in the protein sequence, was shown to be responsible for this enhanced thermostability. Replaceme...
متن کاملDevelopment of a thermostable firefly luciferase
Firefly luciferase forms the basis of a wide range of analytical techniques. However, the enzyme is unstable and rapidly loses activity even at room temperature. This leads to losses in sensitivity and precision in analytical applications and also severely limits the fieldability of devices incorporating luciferase-based technologies. A number of point mutations have previously been identified ...
متن کاملCloning of firefly luciferase cDNA and the expression of active luciferase in Escherichia coli.
A cDNA library was constructed from firefly (Photinus pyralis) lantern poly(A)+ RNA, using the Escherichia coli expression vector lambda gt11. The library was screened with anti-P. pyralis luciferase (Photinus luciferin:oxygen 4-oxidoreductase, EC 1.13.12.7) antibody, and several cDNA clones expressing luciferase antigens were isolated. One clone, lambda Luc1, contained 1.5 kilobase pairs of cD...
متن کاملThermostabilization of firefly luciferase by in vivo directed evolution.
Firefly luciferase is widely used in a number of areas of biotechnology and molecular biology. However, rapid inactivation of wild-type (WT) luciferases at elevated temperatures often hampers their application. A simple non-lethal in vivo screening scheme was used to identify thermostable mutants of luciferase in Escherichia coli colonies. This scheme allowed carrying out each cycle of mutagene...
متن کاملذخیره در منابع من
با ذخیره ی این منبع در منابع من، دسترسی به آن را برای استفاده های بعدی آسان تر کنید
ژورنال
عنوان ژورنال: The FEBS Journal
سال: 2016
ISSN: 1742-464X
DOI: 10.1111/febs.13757