The first activation study of a δ-carbonic anhydrase: TweCAδ from the diatom Thalassiosira weissflogii is effectively activated by amines and amino acids

Figures & data

Figure 1. Amino acids 1–11 and amines 12–19 investigated as TweCAδ activators.

Table 1. Activation of human carbonic anhydrase (hCA) isozymes I, II, and TweCAδ with l-Trp, at 25 °C, for the CO₂ hydration reaction²⁰.

Isozyme	kcat^a	KM^a	(kcat) l-Trp^b	KA^c (μM)
	(s^−1)	(mM)	(s^−1)	l-Trp
hCA I^d	2.0 × 10^5	4.0	3.4 × 10^5	44
hCA II^d	1.4 × 10^6	9.3	4.9 × 10^6	27
TweCAδ^e	1.3 × 10^5	3.9	10.6 × 10^5	0.93

^a 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).

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

^c The activation constant (K_A) for each enzyme was obtained by fitting the observed catalytic enhancements as a function of the activator concentration^21–27. Mean from at least three determinations by a stopped-flow, CO₂ hydrase method²⁰. Standard errors were in the range of 5–10% of the reported values (data not shown).

^d Human recombinant isozymes, from Ref.²⁶.

^e Diatom recombinant enzyme, this work.

Table 2. Activation constants of hCA I, hCA II and the bacterial TweCAδ with amino acids and amines 1–19, by a stopped-flow CO₂ hydrase assay²⁰.

		KA (μM)^a
No.	Compound	hCA I^b	hCA II^b	TweCAδ^c
1	l-His	0.03	10.9	0.75
2	d-His	0.09	43	4.90
3	l-Phe	0.07	0.013	2.15
4	d-Phe	86	0.035	1.96
5	l-DOPA	3.1	11.4	2.11
6	d-DOPA	4.9	7.8	6.24
7	l-Trp	44	27	0.93
8	d-Trp	41	12	0.69
9	l-Tyr	0.02	0.011	1.52
10	d-Tyr	0.04	0.013	0.051
11	4-H2N-l-Phe	0.24	0.15	18.9
12	Histamine	2.1	125	1.34
13	Dopamine	13.5	9.20	0.51
14	Serotonin	45	50	0.90
15	2-Pyridyl-methylamine	26	34	5.28
16	2-(2-Aminoethyl)pyridine	13	15	8.16
17	1-(2-Aminoethyl)-piperazine	7.4	2.30	4.37
18	4-(2-Aminoethyl)-morpholine 0.14	0.19	7.39
19	l-Adrenaline	0.09	96	2.43

^a Mean from three determinations by a stopped-flow, CO₂ hydrase method²⁰. Standard errors were in the range of 5–10% of the reported values (data not shown).

^b Human recombinant isozymes, stopped flow CO₂ hydrase assay method²⁶.

^c Diatom enzyme, this work.

Khalifah RG. The carbon dioxide hydration activity of carbonic anhydrase. I. Stop-flow kinetic studies on the native human isoenzymes B and C. J Biol Chem 1971;246:2561–73.

 PubMed Web of Science ®Google Scholar

(a) Innocenti A, Hilvo M, Parkkila S, et al. Carbonic anhydrase activators. Activation of the membrane-associated isoform XV with amino acids and amines. Bioorg Med Chem Lett 2009;19:3430–3.

 PubMed Web of Science ®Google Scholar

(b) Supuran CT, Dinculescu A, Balaban AT. Carbonic anhydrase activators. Part 5. CA II activation by 2,4,6-trisubstituted pyridinium cations with 1-(ω-aminoalkyl) side chains. Rev Roum Chim 1993;38:343–9.

 Web of Science ®Google Scholar

(c) Supuran CT, Barboiu M, Luca C, et al. Carbonic anhydrase activators. Part 14. Synthesis of mono- and bis- pyridinium salt derivatives of 2-amino-5-(2-aminoethyl)- and 2-amino-5-(3-aminopropyl)-1,3,4-thiadiazole, and their interaction with isozyme II. Eur J Med Chem 1996;31:597–606.

 Web of Science ®Google Scholar

(d) Ilies MA, Banciu MD, Ilies M, et al. Carbonic anhydrase activators. Part 17. Synthesis and activation study of a series of 1-(1,2,4-triazole-(1H)-3-yl)-2,4,6-trisubstituted-pyridinium salts against isozymes I, II and IV. Eur J Med Chem 1997;32:911–8.

 Web of Science ®Google Scholar