TABLE 2.

Phenotype and protein expression

Mutant or revertant type	Strain	Mutation or substitutiona	Location in secondary modelb	Phenotype on melibiose MacConkeyc	Protein expression (%)d
	melB control			White
	Wild-type			Red	100
Site-directed mutante	R52S	Arg-52→Ser	Helix II	White	114 ± 15
	R52V	Arg-52→Val	Helix II	White	101 ± 19
	R52Q	Arg-52→Gln	Helix II	White	112 ± 18
First-site revertantf	R52K	Arg-52→Lys	Helix II	Red	85 ± 12
Second-site revertant	R52S W116R	Trp-116 to Arg	Helix IV	Red	43 ± 8
	R52S S247R	Ser-247 to Arg	Helix VII	Red	105 ± 25
	R52S N248K	Asn-248 to Lys	Helix VII	Red	75 ± 18
	R52S D19G	Asp-19 to Gly	Helix I	Pink	12 ± 3
	R52S D55N	Asp-55 to Asn	Helix II	Red	92 ± 25
	R52S P60Q	Pro-60 to Gln	Helix II	Red	73 ± 3
	R52S N244S	Asn-244 to Ser	Helix VII	Red	72 ± 8
	pR52S I352V	Ile-352 to Val	Loop X-XI	Red	111 ± 21
	R52V T338R	Thr-338 to Arg	Helix X	Red	37 ± 4

^aFor the second-site revertants, only the positions and identities of second-site revertant amino acid substitutions within the melibiose carrier are shown. 

^bPosition of the mutant or revertant amino acid residue within the secondary topological model for the melibiose carrier (Fig. 1). 

^cMacConkey agar is a rich medium lacking a fermentable carbon source. Melibiose was added to the agar as the sole fermentable carbon source. The fermentation of melibiose on this medium causes cells to form red colonies. Cells not able to ferment melibiose form white colonies. The concentration of melibiose was 15 mM (0.5%). 

^dExpression levels for melibiose carriers are reported as percentages of the expression found for the wild-type carrier. The values are averages of at least two separate measurements ± standard errors. 

^eStrains in which Arg-52 has been replaced with Ser, Gln, or Val by site-directed mutagenesis. These strains were subsequently used for the isolation of revertants that regained melibiose transport activity. 

^fFirst-site revertant isolated from the R52Q mutant strain.