NEDD8 covalently binds catalytic cysteine of UBA3:NAE1

NEDD8 is attached via a thioester bond to the catalytic cysteine of its E1 as the first step in its transfer to substrates (Walden et al, 2003). The NEDD8 E1 is a heterodimer consisting of UBA3 and NAE1, and transfers NEDD8 to the E2 enzymes from a 'doubly loaded' state. In the first step, NEDD8 binds to the adenylation site on the NAE1 subunit in conjuction with ATP and Mg2+, generating a covalently modified NEDD8-adenylate conjugate. This conjugation activates the NEDD8 C-terminus for chemical attack by the thiol group of the catalytic cysteine of the UBA3 subunit. Catalysis is likely facilitated by a conformational change in the E1 enzyme. After catalysis, NEDD8 is covalently bound in the E1 catalytic site, leaving the adenylation site free to bind another NEDD8 molecule in the second step, prior to NEDD8 transfer to an E2 enzyme (Walden et al, 2003; Huang et al, 2004; Huang et al, 2005; Huang et al, 2007).

NEDD8通过硫酯键与E1酶的催化半胱氨酸结合，作为其向底物转移的第一步（Walden等，2003）。NEDD8 E1酶由UBA3和NAE1组成的异源二聚体构成，并将NEDD8从“双重负载”状态转移到E2酶。在第一步中，NEDD8与ATP和Mg2+共同作用于NAE1亚基上的腺苷酸化位点，生成共价修饰的NEDD8-腺苷酸共轭物。这种共轭作用激活了NEDD8的C端，使其对UBA3亚基催化半胱氨酸的硫醇基团进行化学攻击。催化过程可能通过E1酶的构象变化而得以促进。催化完成后，NEDD8以共价键的形式结合在E1催化位点上，使得腺苷酸化位点得以空闲，以便在第二步中结合另一个NEDD8分子，在NEDD8转移至E2酶之前（Walden等，2003；Huang等，2004；Huang等，2005；Huang等，2007）。