Srisodsuk M, Reinikainen T, Penttilä M, Teeri T T
VTT, Biotechnical Laboratory, Espoo, Finland.
J Biol Chem. 1993 Oct 5;268(28):20756-61.
Cellobiohydrolase I (CBH I), the major component of Trichoderma reesei cellulolytic system, is comprised of a catalytic core domain joined to a cellulose binding-domain (CBD) by an extended O-glycosylated interdomain linker peptide. Two internal deletions were introduced to the linker in order to investigate its function particularly in the hydrolysis of crystalline cellulose. Deletion of the first one-third of the linker, including a putative hinge region, reduces the binding capacity of CBH I in high enzyme coverage but does not affect its enzymatic activity on crystalline cellulose. The longer deletion removing practically all of the linker dramatically reduces the rate of crystalline cellulose degradation even though the enzyme still binds to the substrate. We conclude that sufficient spatial separation of the two domains is required for efficient function of CBH I. It is evident that the presence of a functional CBD is increasingly important for CBH I toward higher enzyme to cellulose ratios. Our data suggest that the putative hinge removed by the first deletion facilitates CBD-driven binding and dense packing of the wild type enzyme on the cellulose surface.
纤维二糖水解酶I（CBH I）是里氏木霉纤维素分解系统的主要成分，它由一个催化核心结构域和一个纤维素结合结构域（CBD）通过一个延长的O-糖基化结构域间连接肽相连组成。为了特别研究其在结晶纤维素水解中的功能，在连接肽中引入了两个内部缺失。缺失连接肽的前三分之一，包括一个假定的铰链区，在高酶覆盖率下会降低CBH I的结合能力，但不影响其对结晶纤维素的酶活性。更长的缺失几乎去除了整个连接肽，即使酶仍能与底物结合，也会显著降低结晶纤维素的降解速率。我们得出结论，两个结构域之间足够的空间分离是CBH I高效发挥功能所必需的。显然，对于CBH I而言，随着酶与纤维素比例的增加，功能性CBD的存在变得越来越重要。我们的数据表明，第一次缺失所去除的假定铰链促进了野生型酶在纤维素表面的CBD驱动的结合和紧密堆积。
