Covalent Binding of an NAD Analogue to Liver Alcohol Dehydrogenase Resulting in an Enzyme-Coenzyme Complex not Requiring Exogenous Coenzyme for Activity
(1978) In Eur J Biochem 86. p.455-463- Abstract
- 1. The NAD analogue, N6-[N-(6-aminohexyl)carbamoylmethyl]-NAD, was covalently bound to horse liver alcohol dehydrogenase in a carbodiimide-mediated reaction and in such a way that it was active with the very same enzyme molecule to which it was coupled.
2. The degree of substitution, i.e. the number of NAD analogues per enzyme subunit, could be varied (0.3 - 1.6). In one preparation 1.6 coenzyme molecules were bound per subunit; the alcohol dehydrogenase activity of this preparation was 40% of the activity obtained after addition of free NAD in excess.
3. It was calculated that every fourth active site of this preparation was provided with a covalently bound functioning coenzyme analogue, and that this... (More) - 1. The NAD analogue, N6-[N-(6-aminohexyl)carbamoylmethyl]-NAD, was covalently bound to horse liver alcohol dehydrogenase in a carbodiimide-mediated reaction and in such a way that it was active with the very same enzyme molecule to which it was coupled.
2. The degree of substitution, i.e. the number of NAD analogues per enzyme subunit, could be varied (0.3 - 1.6). In one preparation 1.6 coenzyme molecules were bound per subunit; the alcohol dehydrogenase activity of this preparation was 40% of the activity obtained after addition of free NAD in excess.
3. It was calculated that every fourth active site of this preparation was provided with a covalently bound functioning coenzyme analogue, and that this analogue had a cycling rate of about 40000 cycles/h in a coupled substrate assay.
4. The presence of the covalently bound coenzyme made the active sites difficult to inhibit with a competitive inhibitor. For example, 10 mM AMP inhibited the activity of the preparation by 50 % whereas a reference system containing native alcohol dehydrogenase was inhibited by 80 in spite of the fact that the reference system contained about 20000 times as high a concentration of coenzyme. (Less)
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https://lup.lub.lu.se/record/11b4417e-ff89-468d-bd31-f75f23a40cc4
- author
- Månsson, Mats-Olle LU ; Larsson, Per-Olof LU and Mosbach, Klaus LU
- organization
- publishing date
- 1978
- type
- Contribution to journal
- publication status
- published
- subject
- keywords
- Covalently bound NAD analogue, 40 000 cycles/h
- in
- Eur J Biochem
- volume
- 86
- pages
- 9 pages
- publisher
- Wiley-Blackwell
- external identifiers
-
- scopus:0017900445
- ISSN
- 0014-2956
- DOI
- 10.1111/j.1432-1033.1978.tb12328.x
- language
- English
- LU publication?
- yes
- id
- 11b4417e-ff89-468d-bd31-f75f23a40cc4
- date added to LUP
- 2024-06-22 15:28:41
- date last changed
- 2024-09-13 10:17:35
@article{11b4417e-ff89-468d-bd31-f75f23a40cc4,
abstract = {{1. The NAD analogue, N<sup>6</sup>-[N-(6-aminohexyl)carbamoylmethyl]-NAD, was covalently bound to horse liver alcohol dehydrogenase in a carbodiimide-mediated reaction and in such a way that it was active with the very same enzyme molecule to which it was coupled. <br/><br/>2. The degree of substitution, i.e. the number of NAD analogues per enzyme subunit, could be varied (0.3 - 1.6). In one preparation 1.6 coenzyme molecules were bound per subunit; the alcohol dehydrogenase activity of this preparation was 40% of the activity obtained after addition of free NAD in excess.<br/><br/>3. It was calculated that every fourth active site of this preparation was provided with a covalently bound functioning coenzyme analogue, and that this analogue had a cycling rate of about 40000 cycles/h in a coupled substrate assay.<br/><br/>4. The presence of the covalently bound coenzyme made the active sites difficult to inhibit with a competitive inhibitor. For example, 10 mM AMP inhibited the activity of the preparation by 50 % whereas a reference system containing native alcohol dehydrogenase was inhibited by 80 in spite of the fact that the reference system contained about 20000 times as high a concentration of coenzyme.}},
author = {{Månsson, Mats-Olle and Larsson, Per-Olof and Mosbach, Klaus}},
issn = {{0014-2956}},
keywords = {{Covalently bound NAD analogue; 40 000 cycles/h}},
language = {{eng}},
pages = {{455--463}},
publisher = {{Wiley-Blackwell}},
series = {{Eur J Biochem}},
title = {{Covalent Binding of an NAD Analogue to Liver Alcohol Dehydrogenase Resulting in an Enzyme-Coenzyme Complex not Requiring Exogenous Coenzyme for Activity}},
url = {{http://dx.doi.org/10.1111/j.1432-1033.1978.tb12328.x}},
doi = {{10.1111/j.1432-1033.1978.tb12328.x}},
volume = {{86}},
year = {{1978}},
}