Change search
ReferencesLink to record
Permanent link

Direct link
Amidases have a hydrogen bond that facilitates nitrogen inversion but esterases have not
KTH, School of Biotechnology (BIO), Biochemistry.ORCID iD: 0000-0002-4066-2776
KTH, School of Biotechnology (BIO), Biochemistry.
2011 (English)In: ChemCatChem, ISSN 1867-3899, Vol. 3, no 5, 853-860 p.Article in journal (Refereed) Published
Abstract [en]

The fact that proteases/amidases can hydrolyze amides efficiently whereas esterases can not has been discussed during the last decades. By using molecular modeling we have found a hydrogen bond in the transition state for protease/amidase catalyzed hydrolysis of peptides and amides donated by the scissile NH-group of the substrate. The hydrogen-bond acceptor was found either in the enzyme (enzyme assisted) or in the substrate (substrate assisted). This new interaction with the NH-hydrogen in the transition state (TS) was found in sixteen proteases/amidases, which represent ten different reaction mechanisms and eleven different folding families. Esterases lack this interaction and, therefore, they are slow in hydrolyzing amides. By mimicking the substrate-assisted catalysis found in amidases we were able to shift reaction specificity of amide over ester synthesis of Candida antarctica lipase B one hundred fold. We propose that the hydrogen bond facilitates nitrogen inversion in amidases.

Place, publisher, year, edition, pages
2011. Vol. 3, no 5, 853-860 p.
National Category
Biochemistry and Molecular Biology
Identifiers
URN: urn:nbn:se:kth:diva-33330DOI: 10.1002/cctc.201000448ISI: 000290445100008ScopusID: 2-s2.0-80051810428OAI: oai:DiVA.org:kth-33330DiVA: diva2:414465
Note
QC 20110608Available from: 2011-05-03 Created: 2011-05-03 Last updated: 2011-06-08Bibliographically approved
In thesis
1. On electrostatic effects, minimal motion and other catalytic strategies used by enzymes
Open this publication in new window or tab >>On electrostatic effects, minimal motion and other catalytic strategies used by enzymes
2011 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

Enzymes are powerful biocatalysts that provide rate accelerations of up to 1019 fold compared to the corresponding uncatalyzed reaction in solution. The origin of the remarkable performance displayed by enzymes has fascinated and puzzled researchers for over a hundred years. It is clear that the catalytic effect is a consequence of the higher degree of transition state stabilization for the enzyme catalyzed reaction compared to the corresponding uncatalyzed reaction. It is still not well understood exactly how this transition state stabilization occurs and the relative importance of various catalytic effects are discussed. Catalytic effects involving electrostatics, near attack conformers, dynamic effects and an economy in atomic motion are discussed in this thesis.

The importance of electrostatic effects is corroborated in this thesis. A single hydrogen bond in transition state constitutes an important difference between amidases and esterases. A hydrogen bond in transition state is found in all sixteen analyzed amidases representing ten different reaction mechanisms and eleven different folding families. The hydrogen bond is shown to be either substrate assisted or enzyme assisted. The role of this hydrogen bond is to assist nitrogen inversion in amidases. Esterases lack this interaction in transition state and therefore they are very poor catalysts in the hydrolysis of amides. Electrostatic interactions are found to facilitate proton transfer that enhances the rate of lipase catalyzed N-acylation of amino alcohols.

In this thesis electrostatic effects in the substrate are shown to be important for the lipase catalyzed transacylation of acrylates The α,β-double bond present in acrylates introduce electronic effects that has the consequence of restricting the conformational freedom of the substrate in its ground state to two flat conformations, s-cis and s-trans. It is shown that acrylates form near attack conformers (NACs) from their ground state s-cis/s-trans planar conformations. The ability of the enzyme to accommodate such apparent s-cis/s-trans substrate conformations dictates the probability to form productive transition states and thus the reaction rate.

Dynamic effects are important in enzymes. In this thesis it is found that a point mutation increases the flexibility of a neighbouring residue in Candida antarctica lipase B. This allows the mutated enzyme to explore conformations not accessible for the wild-type enzyme. The dynamics has the effect to decrease steric interactions in transition state with concomitant rate increase for the transacylation of methyl methacrylate.

In this thesis an economy of atomic motion during enzyme catalysis is observed. Nitrogen inversion in amidases constitutes an interesting example. A rotation as part of the reaction mechanism for amide bond hydrolysis would involve much more motion.

Place, publisher, year, edition, pages
Stockholm: KTH Royal Institute of Technology, 2011. 42 p.
Series
Trita-BIO-Report, ISSN 1654-2312 ; 14
National Category
Biochemistry and Molecular Biology
Research subject
SRA - Molecular Bioscience
Identifiers
urn:nbn:se:kth:diva-33334 (URN)978-91-7415-976-9 (ISBN)
Public defence
2011-05-27, D2, Lindstedtsvägen 5, Stockholm, 13:00 (English)
Opponent
Supervisors
Note
QC 20110512Available from: 2011-05-12 Created: 2011-05-03 Last updated: 2011-05-12Bibliographically approved

Open Access in DiVA

No full text

Other links

Publisher's full textScopus

Search in DiVA

By author/editor
Syrén, Per-OlofHult, Karl
By organisation
Biochemistry
Biochemistry and Molecular Biology

Search outside of DiVA

GoogleGoogle Scholar

Altmetric score

Total: 107 hits
ReferencesLink to record
Permanent link

Direct link