. 1998 Jun;42(6):1375–1381. doi: 10.1128/aac.42.6.1375

TABLE 3.

Kinetic characteristics of recombinant Y49 β-lactamase from E. coli and β-lactamase obtained directly from M. tuberculosis

Substrate	Kinetics of hydrolysis^a
	Recombinant Y-49 β-lactamase					Directly from M. tuberculosis
	Recombinant Y-49 β-lactamase					Post-anion-exchange fraction			Chromatofocusing peak 2 fraction
	K_m (μM)	k_cat (s⁻¹)	Relative V_max^b	REH^c	K_i (μM)	K_m (μM)	Relative V_max^b	REH^c	K_m (μM)	Relative V_max^b	REH^c
Benzylpenicillin	50	21	100	100		58	100	100	74	100	100
Phenoxymethylpenicillin	38	20	95	125					78	70	66
Amoxicillin	94	2	10	5		95	17	10
Azlocillin	185	55	262	71		340	363	62	326	338	77
Nitrocefin	81	31	147	91		114	180	92	105	185	131
Cephaloridine	798	8	39	2		540	23	2
Cefazolin	490	8	36	4		304	17	3
Cephalothin	308	8	36	6		218	25	7
Cephapirin	680	3	15	1		319	9	2
Cefamandole	645	22	104	8		282	63	13	566	126	16
Cefuroxime^d							0.4
Ceftriaxone^d							<0.5
Clavulanic acid					0.4
Sulbactam					0.5

Values for the recombinant Y49 β-lactamase and the pooled post-DEAE-cellulose anion-exchange chromatography fractions represent the averages of three or four assays of rates of hydrolysis of substrates. Values for the chromatofocusing peak 2, i.e., the pI 4.9 enzyme, were obtained from a single experiment.

V_max values are relative to the V_max of benzylpenicillin, which was assigned a value of 100. The measured V_max values for benzylpenicillin were 41, 0.31, and 0.51 μmol hydrolyzed per min per mg of protein, respectively, for the purified recombinant Y49 β-lactamase, the pooled post-anion-exchange fractions (containing both the pI 5.1 and the pI 4.9 β-lactamases), and the peak 2 fraction (containing the pI 4.9 β-lactamase) obtained following chromatofocusing.

Relative efficiency of hydrolysis (REH) values are derived from the V_max/K_m ratio, with the benzylpenicillin value adjusted to 100. The other relative efficiency of hydrolysis values are expressed as a percentage of the value for benzylpenicillin.

The rates of degradation of cefuroxime and ceftriaxone were too low to obtain K_m and V_max values. For these β-lactams, the initial velocity of hydrolysis of a 100 μM solution of the cephalosporin is given as a percentage of the initial velocity of hydrolysis of a 1,000 μM solution of benzylpenicillin.