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Publication Abstract from PubMed

Thermostable enzymes have significant advantages in industries, yet uncovering novel candidates with superior properties remains a scientific pursuit. This study identified rMxyl(cd), a glycoside hydrolase 11 family thermophilic xylanase from compost-soil metagenome, which exhibited a high specific activity of 5954 U.mg(-1) at pH 5.5 and 80 degrees C. rMxyl(cd) was crystallized and diffracted to 1.5 A resolution. Compared to the mesophilic xylanase Xyn II, rMxyl(cd) exhibits a more compact architecture. Notably, B-factor analysis reveals a uniquely flexible thumb region, hinting at its critical role in the enzyme's catalytic mechanism. rMxyl(cd) contains two disulfide bonds in the thumb and the N-terminal regions. Breaking these disulfide bonds by mutagenesis has dramatically decreased activities and thermostability. Conversely, introducing an extra disulfide bond at the N-terminal region of its alpha-helix extended its half-life for more than five folds at 80 degrees C. Our studies firmly establish that the disulfide bonds are essential for its high thermal stability and the flexibility of the thumb region is crucial for its activity. Comparing the rMxyl(cd) crystal structure with the AlphaFold2-predicted model shows overall similarity, but the crystal structure offers higher local accuracy, especially in key functional regions. These findings not only deepen our understanding of the structure-function relationship of thermophilic xylanases but also inform a rational design of industrial enzymes.

, PMID:40368259^[1]

From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine.