Table 1.

Properties of spin-labeled cysteine mutants of LacY

Spin-labeled mutants	Nitroxide positions	NPGal binding* (relative to unlabeled mutant), %	Distances modeled for apo LacY†
			Ca-Ca, Å	Spin – spin distances, Å
				Short	Long	Average
R73C/S401C/WT	Cytoplasmic ends of helices III and XII	69	41	44	52	48
R73C/S401C/C154G		93
R73C/Q340C/WT	Cytoplasmic ends of helices III and X	63	36	38	48	43
R73C/Q340C/C154G		73
S136C/Q340C/WT	Cytoplasmic ends of helices IV and X	62	34	39	51	45
N137C/Q340C/WT	Cytoplasmic ends of helices IV and X	95	32	30	46	38
S136C /S401C/WT	Cytoplasmic ends of helices IV and XII	60	40	43	55	49
N137C /S401C/WT	Cytoplasmic ends of helices IV and XII	57	38	33	52	43
V105C/T310C/WT	Periplasmic ends of helices IV and IX	80	34	30	50	40
V105C/T310C/C154G		94
I164C/T310C/WT	Periplasmic ends of helices V and IX	97	27	24	41	33
I164C/T310C/C154G		88
I164C/S375C/WT	Periplasmic ends of helices V and XI	94	33	30	45	38

*Detected by direct sugar-binding assay based on Trp-NPGal FRET as described in SI Materials and Methods.

^†Estimated using LacY crystal structure (PDB ID: 1PV6) as described in Materials and Methods.