Table 1.

Effects of substitutions on subunit interactions and viral replication

M50 (1–168) mutation	Kd (µM)*	Factor weaker than WT†	UL50 (1–169) mutation	Kd (µM)	Factor weaker than WT	HCMV replication phenotype‡
WT	0.97 ± 0.12	1	WT	0.29	1	WT
V52A	NB	>100	V52A	1.4 ± 0.72	5	5- to 10-fold lower yield
E56A	N/A	N/A	E56A	30–90	>100	Nonviable
Y57A	4.1 ± 0.35	5	Y57A	NB	>100	N/A
S60A	17	20	S60A	N/A	N/A	N/A
Y61A	Insoluble	N/A	Y61A	Insoluble	N/A	N/A
			E63A	1.1 ± 0.33	4	N/A
			R65A	0.55 ± 0.08	2	N/A
I121A	98	>100	I122A	N/A	N/A
C124A	76	87	S125A	2.2	7	5- to 10-fold lower yield
V128A	0.89	1	V129A	0.32	1	N/A
L129A	60–85	68–100	L130A	7.9	27	Nonviable
			C133A	0.67 ± 0.30	1	N/A
D153A	0.34 ± 0.14	0.4	E154A	0.51 ± 0.24	2	N/A
N154A	16 ± 2.5	16	N155A	N/A	N/A	N/A

N/A, not generated or tested (the M50 and UL50 Y61A mutant proteins were insoluble); NB, no detectable binding.

^*K_D values ± SD were derived from ITC data for binding of M53 (residues 103–333) to WT and single point mutant M50 (residues 1–168) proteins and binding of UL53 (residues 50–292) to WT and single point mutant UL50 (residues 1–169) proteins.

^†Factors weaker than WT values were calculated by dividing the K_d value of the mutant by the K_d value of the WT protein.

^‡Mutant BACs that produced no detectable spread or virus were considered nonviable. Factor reductions in yield of viable mutants were calculated by dividing the supernatant viral titer of the mutant virus by the supernatant viral titer of WT virus at day 7 postinfection (Fig. S5A).