. 2015 Jun 22;290(32):19584–19596. doi: 10.1074/jbc.M115.661371

TABLE 2.

Substrate(s) in active site formed by the cyclase dimer

Conversion of the substrate highlighted in bold to the respective cyclic monophosphate was tested (Fig. 10) to probe for the presence of the corresponding functional cyclase dimer.

RetGC1 dimer (subunit A/subunit B)	Substrate in active site (subunit A/subunit B)
mOrangeRetGC1/mOrangeRetGC1^a	GTP/GTP^a
GC1AC/GC1AC^b	ATP/ATP^b
GC1AC rib(−)/GC1AC rib(−)^c	Inactive/inactive^c
mOrangeM823R/mOrangeM823R^a	GTP/GTP^a
mOrangeR822P/mOrangeR822P^a	GTP/GTP^a
mOrangeM823R/GC1AC rib(−)^d	Inactive/ATP^d^,^e
mOrangeR822P/GC1AC rib(−)^d	Inactive/ATP^d^,^e
mOrangeM823R/GC1AC^f	GTP/ATP^f
mOrangeRetGC1/GC1AC rib(−)^d	Inactive/ATP^d^,^e

^a Each subunit of RetGC1 in a dimer coordinates the purine base of GTP and provides coordination of ribose and Mg²⁺ for the second Mg²⁺GTP molecule in the active site (Fig. 10A) (28).

^b RetGC1 with short ECD (37) was converted to adenylyl cyclase by the E925K/C997D substitutions as in Ref. 44.

^c In addition to the E925K/C997D substitutions, the GC1AC also had the D929A mutation (24), disabling coordination of ribose and eliminating catalytic activity.

^d The purine base of GTP is recognized by subunit A, but the opposite subunit B fails to coordinate ribose of the GTP, whereas ATP bound through the purine base by subunit B is held in a normal fashion by the Asp⁹²⁹ of subunit A (28).

^e Subunit A cannot utilize Mg²⁺GTP as a substrate because subunit B cannot coordinate the ribose and Mg²⁺ of GTP; subunit B coordinates the purine base of ATP, whereas subunit A provides coordination of ribose and Mg²⁺ of the ATP (24, 28, 44).

^f Subunit A coordinates purine base of GTP, and subunit B coordinates that of ATP; each subunit provides coordination of ribose and Mg²⁺ for the NTP whose purine base is recognized by the opposite subunit (28, 44).