Rational and Irrational Approaches to Convince a Protein to Crystallize

Abts, André; Schwarz, Christian; Tschapek, Britta; Smits, Sander H. J.; Schmitt, Lutz

doi:10.5772/28014

Cited by 2 publications

(2 citation statements)

References 127 publications

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“…Beyond that all requirements for the protein solution itself remain valid as described in [1] in more detail.…”

Section: General Approaches To Obtain Crystals With Bound Ligands And...mentioning

confidence: 99%

“…Crystallization of a protein is a tedious route and although a lot of knowledge about crystallization has been gained in the last decades, one still cannot predict the outcome. The sometimes unexpected bottlenecks in protein purification and crystallization have recently been summarized and possible strategies to obtain a protein crystal were postulated [1]. This book chapter will tackle the next step: How to crystallize protein-ligand complexes or intermediate steps of the reaction cycle?…”

Section: Introductionmentioning

confidence: 99%

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Proteins and Their Ligands: Their Importance and How to Crystallize Them

Hoeppner¹,

Schmitt²,

Smits

2013

Advanced Topics on Crystal Growth

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usage of catalytically inactive mutants with bound ligand(s)• creation of an environment (i. g. buffer condition) which shifts the equilibrium constant so that the reaction cannot occurThe most important point concerning preparing co-crystallization trials is the knowledge of the corresponding kinetical parameters. Proteins bind their natural ligand(s) with high affinity, which means in the nM-up to low mM range. To successfully crystallize a protein with the ligand(s) bound, the affinity needs to be determined. There are numerous biophysical techniques to achieve this, for example Intrinsic Tryptophan Fluorescence, Isothermal Calorimetry, Surface Plasmon Resonance and many others. In principle the affinity is determined by the size of a ligand as well as the property of the binding site of the protein. As first approximation, one can state that affinity increases with a decrease in ligand size.

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“…Beyond that all requirements for the protein solution itself remain valid as described in [1] in more detail.…”

Section: General Approaches To Obtain Crystals With Bound Ligands And...mentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Proteins and Their Ligands: Their Importance and How to Crystallize Them

Hoeppner¹,

Schmitt²,

Smits

2013

Advanced Topics on Crystal Growth

Self Cite

View full text Add to dashboard Cite

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Time-Dependent Protein Thermostability Assay

Vandecaetsbeek

Vangheluwe

2016

P-Type ATPases

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Membrane protein purification often yields rather unstable proteins impeding functional and structural protein characterization. Low protein stability also leads to low purification yields as a result of protein degradation, aggregation, precipitation, and folding instability. It is often required to optimize buffer conditions through numerous iterations of trial and error to improve the homogeneity, stability, and solubility of the protein sample demanding high amounts of purified protein. Therefore we have set up a fast, simple, and high-throughput time-dependent thermostability-based assay at low protein cost to identify protein stabilizing factors to facilitate the handling and characterization of membrane proteins by subsequent structural and functional studies.

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Rational and Irrational Approaches to Convince a Protein to Crystallize

Cited by 2 publications

References 127 publications

Proteins and Their Ligands: Their Importance and How to Crystallize Them

Proteins and Their Ligands: Their Importance and How to Crystallize Them

Time-Dependent Protein Thermostability Assay

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