Cryo-EM structures of HKU2 and SADS-CoV spike glycoproteins and insights into coronavirus evolution

Jinfang Yu; Shuyuan Qiao; Runyu Guo; Xinquan Wang

Two of them (C 590 -C 612 and C 595 -C 601 ) stabilize the folded helices of UH, C 696 -C 706 308 bends the CR, C 884 -C 895 links the CH and the BH, and C 934 -C 943 is within the SD3 (Fig. 309 6a ). The first four disulfide bonds are conserved in all coronaviruses, and the last one Fig. 4) . The typical CR 321 in the S2 subunit contains three helices and one short strand, with a disulfide bond 322 bending the first and second helix to form a turning (Fig. 6b) . In HKU2, the second 323 helix is replaced by a short strand (713-716) and the third helix is replaced by a loop 324 (721-741), therefore there are two short strands and only one helix in HKU2 CR (Fig. 325 6c). The conserved disulfide bond C 696 -C 706 makes the first helix of CR in HKU2 spike 326 turn upside down. The S2' cleavage site (between R671 and S672) is then covered by 327 the reversed CR helix and loops, and R671 interacts with E723 in the loop and K697 328 and K698 in helix 1 (Fig. 6c) . In other coronaviruses, taking the MHV S2 for example, 329 the helix 1 does not cover the S2' site (between R869 and S870), and R869 only loosely 330 interacts with T929 in helix 3 (Fig. 6b) . After the dissociation of the S1 subunit triggered 331 by receptor binding, the exposure of the S2' site for cleavage is a prerequisite for the 332 proteolytic activation of the coronavirus spike to mediate membrane fusion. The buried 333 author/funder. All rights reserved. No reuse allowed without permission.