Table 2.

Point mutation analyses of known PhK/PKA phosphorylatable serines within the 32 residue N-terminus of the PhK β subunit.

	AD	Vector encoding pB42
BD		βFL	βS11AS26A	βS11AS26E	βS11ES26A	βS11ES26E	Empty vector
Vector encoding pLexA	βFL	2.93 ± 0.04	3.14 ± 0.06	7.98 ± 0.04	10.48 ± 0.06	6.81 ± 0.05	2.06 ± 0.07
	β32N	8.36 ± 0.14	1.79 ± 0.02	18.95 ± 0.17	14.96 ± 0.06	9.79 ± 0.06	3.16 ± 0.07
	βS11A	6.51 ± 0.12	1.39 ± 0.05	6.33 ± 0.04	8.09 ± 0.06	6.37 ± 0.06	2.30 ± 0.08
	βS11E	5.04 ± 0.07	0.67 ± 0.07	2.15 ± 0.03	7.07 ± 0.06	3.86 ± 0.07	1.41 ± 0.04
	βS26A	6.47 ± 0.07	2.53 ± 0.06	3.51 ± 0.06	12.20 ± 4.96	5.83 ± 0.06	3.36 ± 0.06
	βS26E	5.82 ± 0.07	3.21 ± 0.03	5.66 ± 0.04	12.40 ± 0.07	4.40 ± 0.02	3.54 ± 0.06
	βS11AS26A	5.98 ± 0.07	1.81 ± 0.05	6.14 ± 0.04	14.89 ± 0.07	5.04 ± 0.03	2.90 ± 0.03
	βS11AS26E	4.78 ± 0.08	1.13 ± 0.04	9.01 ± 0.06	13.61 ± 0.12	1.85 ± 0.03	2.05 ± 0.07
	βS11ES26A	5.47 ± 0.14	1.00 ± 0.05	7.06 ± 0.02	7.86 ± 0.07	3.86 ± 0.05	2.90 ± 0.06
	βS11ES26E	6.48 ± 0.12	1.07 ± 0.05	10.20 ± 0.07	13.99 ± 0.11	4.22 ± 0.04	2.89 ± 0.06
	Empty vector	0.87 ± 0.02	0.77 ± 0.04	0.58 ± 0.04	0.82 ± 0.03	0.52 ± 0.05	1.22 ± 0.05

The chemical structure and charge of phosphoserine side-chains are approximated by Ser/Gln mutations. Ser/Ala mutations serve as negative controls for amino acid substitutions at the indicated positions in the primary sequence of β. β-Galactosidase activity from yeast lysates was determined using the substrate β-o-D-galactopyranoside as described under Experimental Procedures. Activity is expressed in units according to the formula: 1000*[(A_420t1 - A_420t2)/(A₆₀₀*Δ_tA₄₂₀)]. Data represent the mean ± S.E. of eight assays. A positive interaction is determined as being significantly greater than the maximum observed control. AD and BD indicate activation and binding domains.