Hult Eva-Lena, Katouno Fuminori, Uchiyama Taku, Watanabe Takeshi, Sugiyama Junji
Research Institute for Sustainable Humanosphere, Kyoto University, Uji, Kyoto 611-0011, Japan.
Biochem J. 2005 Jun 15;388(Pt 3):851-6. doi: 10.1042/BJ20050090.
Beta-chitin microfibrils were treated with ChiA and ChiB (chitinases A and B respectively) from Serratia marcescens 2170. The beta-chitin microfibrils were shortened, and the tips appeared narrowed and sharpened at both ends, after either consecutive or simultaneous degradation by ChiA and ChiB. Increased production of reducing sugars by simultaneous degradation (by ChiA and ChiB) of beta-chitin, but not of glycol chitin, suggests synergistic interactions between the two enzymes. A combined analysis using the tilt microdiffraction method to determine the crystallographic axes, together with the biotin-streptavidin-gold-labelling method specific to the reducing ends, was used to investigate the polarity of the degraded beta-chitin microcrystals. The digestion of the beta-chitin fibrils by ChiA occurred from the reducing end to the nonreducing end, whereas digestion by ChiB occurred from the non-reducing end to the reducing end. The results are in agreement with the previously determined three-dimensional structures of these enzymes.
用来自粘质沙雷氏菌2170的ChiA和ChiB（分别为几丁质酶A和B）处理β-几丁质微纤维。在被ChiA和ChiB连续或同时降解后，β-几丁质微纤维变短，并且其两端变窄且变尖。通过β-几丁质（而非乙二醇几丁质）被ChiA和ChiB同时降解增加了还原糖的产量，这表明这两种酶之间存在协同相互作用。使用倾斜微衍射方法确定晶体学轴，并结合还原端特异性的生物素-链霉亲和素-金标记方法进行联合分析，以研究降解的β-几丁质微晶的极性。ChiA对β-几丁质纤维的消化从还原端向非还原端进行，而ChiB的消化则从非还原端向还原端进行。结果与先前确定的这些酶的三维结构一致。
