TABLE 2.

Initial turnover rates of the glycosylation reaction of peptides with different sequons

Turnover rates are depicted as number of peptide that each molecule of ApNGT is turning over per minute. For determination of relative turnover, the fastest turnover rate (5CF-GSDQNATF) was defined as 100% and the other turnover rates are given in relationship to that rate. Parameters were determined from the kinetic data represented in Fig. 4. Unnatural amino acids: hS, homoserine; Dab, diaminobutyric acid; N(CH₃), N-Methyl-asparagine (side chain structures are depicted in supplemental Fig. S1B).

Peptide sequence	Glycopeptide detected	Initial turnover rate	Relative turnover
		peptide/min	%
Sequon numbering	5CF-G−4S-3D−2Q−1X0A+1Y+2F+3
Variation at position +2
5CF-GSDQNATF	Yes	0.04 ± 0.001	100
5CF-GSDQNASF	Yes	2.9 × 10−3 ± 2.7 × 10−4	7.9
5CF-GSDQNAAF	Yes	3.6 × 10−6 ± 5.7 × 10−7	0.01
5CF-GSDQNAVF	Yes	4.5 × 10−6 ± 5.6 × 10−7	0.01
Variation at position 0
5CF-GSDQQATF	Yes	2.2 × 10−6 ± 3.8 × 10−7	0.006
5CF-GSDQhSATF	Yes	3.3 × 10−4 ± 1.5 × 10−5	0.9
5CF-GSDQDATF	No	NDa	ND
5CF-GSDQDabATF	No	ND	ND
5CF-GSDQN(CH3)ATF	No	ND	ND

^a ND, not determined.