The failure of polypeptides to achieve conformational maturation following biosynthesis can result in the formation of protein aggregates capable of disrupting essential cellular functions. In the secretory pathway, misfolded asparagine (N)-linked glycoproteins are selectively sorted for endoplasmic reticulum-associated degradation (ERAD) in response to the catalytic removal of terminal alpha-linked mannose units. Remarkably, ER mannosidase I/Man1b1, the first alpha-mannosidase implicated in this conventional N-glycan-mediated process, can also contribute to ERAD in an unconventional, catalysis-independent manner. To interrogate this functional dichotomy, the intracellular fates of two naturally occurring misfolded N-glycosylated variants of human alpha1-antitrypsin (AAT), Null Hong Kong (NHK), and Z (ATZ), in Man1b1 knockout HEK293T cells were monitored in response to mutated or truncated forms of transfected Man1b1. As expected, the conventional catalytic system requires an intact active site in the Man1b1 luminal domain. In contrast, the unconventional system is under the control of an evolutionarily extended N-terminal cytoplasmic tail. Also, N-glycans attached to misfolded AAT are not required for accelerated degradation mediated by the unconventional system, further demonstrating its catalysis-independent nature. We also established that both systems accelerate the proteasomal degradation of NHK in metabolic pulse-chase labeling studies. Taken together, these results have identified the previously unrecognized regulatory capacity of the Man1b1 cytoplasmic tail and provided insight into the functional dichotomy of Man1b1 as a component in the mammalian proteostasis network.

多肽在生物合成后无法实现构象成熟的原因可能导致形成能够破坏基本细胞功能的蛋白质聚集体。在分泌途径中，响应末端α-连接的甘露糖单元的催化去除，选择性折叠错误折叠的天冬酰胺（N）-连接的糖蛋白进行内质网相关降解（ERAD）。值得注意的是，ER甘露糖苷酶I / Man1b1是这种常规N-聚糖介导的过程中涉及的第一个α-甘露糖苷酶，也可以以非常规的，不依赖于催化的方式参与ERAD。为了询问这种功能性的二分法，人类自然存在的两种错误折叠的人类α1-抗胰蛋白酶（AAT）的N-糖基化变体，Null Hong Kong（NHK）和Z（ATZ）在Man1b1基因敲除中，对突变的或截短形式的转染Man1b1基因的HEK293T细胞进行了监测。如预期的那样，常规催化系统需要Man1b1腔域中完整的活性位点。相反，非常规系统处于进化延伸的N末端细胞质尾巴的控制之下。而且，由非常规系统介导的加速降解也不需要附着在错误折叠的AAT上的N-聚糖，这进一步证明了其催化独立性。我们还确定，在代谢脉冲追踪标记研究中，两种系统均会加速NHK的蛋白酶体降解。在一起