Comparison of the amine/amino acid activation profiles of the β- and γ-carbonic anhydrases from the pathogenic bacterium Burkholderia pseudomallei

Figures & data

Figure 1. Chemical structures of compounds 1–19 investigated as CAAs in the present paper.

Table 1. Activation of human carbonic anhydrase (hCA) isozymes I, II and BpsCAγ/β with L-Tyr, at 25 °C, for the CO2 hydration reaction⁵³.

	kcat^d	KM^d	(kcat)L-Tyr^e	KA^f (μM)
Isozyme	(s^−1)	(mM)	(s^−1)	L-Tyr
hCA I^a	2.0 × 10^5	4.0	13.9 × 10^5	0.020
hCA II^a	1.4 × 10^6	9.3	12.8 × 10^6	0.011
BpsCAγ^b	5.3 × 10^5	21.2	13.8 × 10^5	0.200
BpsCAβ^c	1.6 × 10^5	4.7	3.10 × 10^6	0.003

^a Human recombinant isozymes, from Ref.⁷.

^b Bacterial recombinant enzyme, from Ref.⁵².

^c Bacterial recombinant enzyme, this work.

^d Observed catalytic rate without activator. K_M values in the presence and the absence of activators were the same for the various CAs (data not shown).

^e Observed catalytic rate in the presence of 10 μM activator.

^f The activation constant (K_A) for each enzyme was obtained by fitting the observed catalytic enhancements as a function of the activator concentration⁵³. Mean from at least three determinations by a stopped-flow, CO₂ hydrase method. Standard errors were in the range of 5–10% of the reported values (data not shown).

Table 2. Activation constants of hCA I, hCA II and the bacterial CAs BpsCAγ/β with amino acids and amines 1–19. Data for hCA I, II and BpsCAγ are from Refs.^25,52.

			KA (μM)^d
No.	Compound	hCA I^a	hCA II^a	BpsγCA^b	BpsCAβ^c
1	L-His	0.03	10.9	24.7	31.6
2	D-His	0.09	43	0.086	0.98
3	L-Phe	0.07	0.013	1.73	3.42
4	D-Phe	86	0.035	0.13	0.075
5	L-DOPA	3.1	11.4	0.072	0.009
6	D-DOPA	4.9	7.8	0.98	0.007
7	L-Trp	44	27	0.43	0.002
8	D-Trp	41	12	0.052	0.001
9	L-Tyr	0.02	0.011	0.20	0.003
10	D-Tyr	nt	nt	32.8	1.89
11	4-H2N-L-Phe	0.24	0.15	0.009	0.0009
12	Histamine	2.1	125	0.12	0.012
13	Dopamine	13.5	9.2	0.014	0.006
14	Serotonin	45	50	0.10	0.027
15	2-Pyridyl-methylamine	26	34	2.36	0.016
16	2-(2-Aminoethyl)pyridine	13	15	0.034	0.94
17	1-(2-Aminoethyl)-piperazine	7.4	2.3	0.018	0.004
18	4-(2-Aminoethyl)-morpholine 0.14	0.19	0.015	0.073
19	L-Adrenaline	0.09	96	0.019	0.002

^a Human recombinant isozymes, stopped flow CO₂ hydrase assay method²⁵.

^b From Ref.⁵², stopped flow CO₂ hydrase assay method.

^c This work.

^d Mean from three determinations by a stopped-flow, CO₂ hydrase method¹⁴. Standard errors were in the range of 5–10% of the reported values (data not shown); nt: not tested.

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