Not Cleaving the His-tag of Thal Results in More Tightly Packed and Better-Diffracting Crystals

Not Cleaving the His-tag of Thal Results in More Tightly Packed and Better-Diffracting Crystals

Flavin-dependent halogenases chlorinate or brominate their substrates in an environmentally friendly manner, only requiring the cofactor reduced flavin adenine dinucleotide (FADH<sub>2</sub>), oxygen, and halide salts. The tryptophan 6-halogenase Thal exhibits two flexible loops, which b...

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Bibliographic Details
Main Authors: Ann-Christin Moritzer, Tina Prior, Hartmut H. Niemann
Format: Article
Language:English
Published: MDPI AG 2020-12-01
Series:Crystals
Subjects:
flavin-dependent halogenase
protein crystallization
tryptophan halogenase
ThdH
Online Access:https://www.mdpi.com/2073-4352/10/12/1135
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spelling doaj-68a2b5bd29d543478155334b2dbb04a32020-12-13T00:01:18ZengMDPI AGCrystals2073-43522020-12-01101135113510.3390/cryst10121135Not Cleaving the His-tag of Thal Results in More Tightly Packed and Better-Diffracting CrystalsAnn-Christin Moritzer0Tina Prior1Hartmut H. Niemann2Structural Biochemistry, Department of Chemistry, Bielefeld University, 33615 Bielefeld, GermanyStructural Biochemistry, Department of Chemistry, Bielefeld University, 33615 Bielefeld, GermanyStructural Biochemistry, Department of Chemistry, Bielefeld University, 33615 Bielefeld, GermanyFlavin-dependent halogenases chlorinate or brominate their substrates in an environmentally friendly manner, only requiring the cofactor reduced flavin adenine dinucleotide (FADH<sub>2</sub>), oxygen, and halide salts. The tryptophan 6-halogenase Thal exhibits two flexible loops, which become ordered (substrate-binding loop) or adopt a closed conformation (FAD loop) upon substrate or cofactor binding. Here, we describe the structure of N<sub>His</sub>-Thal-RebH5 containing an N-terminal His-tag from pET28a, which crystallized in a different space group (<i>P</i>2<sub>1</sub>) and, surprisingly, diffracted to a higher resolution of 1.63 Å than previously deposited Thal structures (<i>P</i>6<sub>4</sub>; ~2.2 Å) with cleaved His-tag. Interestingly, the binding of glycine in the active site can induce an ordered conformation of the substrate-binding loop.https://www.mdpi.com/2073-4352/10/12/1135flavin-dependent halogenaseprotein crystallizationtryptophan halogenaseThdH
collection DOAJ
language English
format Article
sources DOAJ
author Ann-Christin Moritzer
Tina Prior
Hartmut H. Niemann
spellingShingle Ann-Christin Moritzer
Tina Prior
Hartmut H. Niemann
Not Cleaving the His-tag of Thal Results in More Tightly Packed and Better-Diffracting Crystals
Crystals
flavin-dependent halogenase
protein crystallization
tryptophan halogenase
ThdH
author_facet Ann-Christin Moritzer
Tina Prior
Hartmut H. Niemann
author_sort Ann-Christin Moritzer
title Not Cleaving the His-tag of Thal Results in More Tightly Packed and Better-Diffracting Crystals
title_short Not Cleaving the His-tag of Thal Results in More Tightly Packed and Better-Diffracting Crystals
title_full Not Cleaving the His-tag of Thal Results in More Tightly Packed and Better-Diffracting Crystals
title_fullStr Not Cleaving the His-tag of Thal Results in More Tightly Packed and Better-Diffracting Crystals
title_full_unstemmed Not Cleaving the His-tag of Thal Results in More Tightly Packed and Better-Diffracting Crystals
title_sort not cleaving the his-tag of thal results in more tightly packed and better-diffracting crystals
publisher MDPI AG
series Crystals
issn 2073-4352
publishDate 2020-12-01
description Flavin-dependent halogenases chlorinate or brominate their substrates in an environmentally friendly manner, only requiring the cofactor reduced flavin adenine dinucleotide (FADH<sub>2</sub>), oxygen, and halide salts. The tryptophan 6-halogenase Thal exhibits two flexible loops, which become ordered (substrate-binding loop) or adopt a closed conformation (FAD loop) upon substrate or cofactor binding. Here, we describe the structure of N<sub>His</sub>-Thal-RebH5 containing an N-terminal His-tag from pET28a, which crystallized in a different space group (<i>P</i>2<sub>1</sub>) and, surprisingly, diffracted to a higher resolution of 1.63 Å than previously deposited Thal structures (<i>P</i>6<sub>4</sub>; ~2.2 Å) with cleaved His-tag. Interestingly, the binding of glycine in the active site can induce an ordered conformation of the substrate-binding loop.
topic flavin-dependent halogenase
protein crystallization
tryptophan halogenase
ThdH
url https://www.mdpi.com/2073-4352/10/12/1135
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