Efficient protein crystallization

DeLucas, Lawrence J.; Bray, Terry; Nagy, Lisa; McCombs, Debbie; Chernov, N.; Hamrick, David; Cosenza, Larry; Belgovskiy, Alexander; Stoops, Brad; Chait, Arnon

doi:10.1016/s1047-8477(03)00050-9

Cited by 71 publications

(48 citation statements)

References 19 publications

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“…Robotic systems have the advantage of exceptional sample-record maintenance, most can deploy submicrolitre amounts of mother liquor and they can be used to screen vast matrices of conditions that might otherwise be impossible in a practical sense for a lone investigator. Robotic systems are, in addition, now being used to examine and evaluate the results of crystallization trials using optical subsystems and image-processing techniques (Hosfield et al, 2003;DeLucas et al, 2003;Luft et al, 2003). Evaluation of trial arrays of conditions, however, continues to be problematic because of the continuing difficulty in devising meaningful scoring criteria in the absence of actual crystals.…”

Section: Figurementioning

confidence: 99%

“…In recent years, robots and other automated instruments, and entire integrated systems, have been developed to accelerate the crystallization process (Bard et al, 2004;DeLucas et al, 2003;Hosfield et al, 2003;Luft et al, 2003). They have the capacity to screen thousands of crystallization conditions, and they do so precisely and reliably, with fewer errors and better record keeping An observation of inadvertent heterogeneous nucleation of protein crystals that is not uncommon is that of crystals growing along the length of a cotton fiber present in the mother liquor.…”

Section: Robotics and Automation Of Crystallization Setups And Their mentioning

confidence: 99%

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Introduction to protein crystallization

McPherson

Gavira

2013

Acta Crystallogr F Struct Biol Commun

357

323

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Protein crystallization was discovered by chance about 150 years ago and was developed in the late 19th century as a powerful purification tool and as a demonstration of chemical purity. The crystallization of proteins, nucleic acids and large biological complexes, such as viruses, depends on the creation of a solution that is supersaturated in the macromolecule but exhibits conditions that do not significantly perturb its natural state. Supersaturation is produced through the addition of mild precipitating agents such as neutral salts or polymers, and by the manipulation of various parameters that include temperature, ionic strength and pH. Also important in the crystallization process are factors that can affect the structural state of the macromolecule, such as metal ions, inhibitors, cofactors or other conventional small molecules. A variety of approaches have been developed that combine the spectrum of factors that effect and promote crystallization, and among the most widely used are vapor diffusion, dialysis, batch and liquid-liquid diffusion. Successes in macromolecular crystallization have multiplied rapidly in recent years owing to the advent of practical, easy-to-use screening kits and the application of laboratory robotics. A brief review will be given here of the most popular methods, some guiding principles and an overview of current technologies.

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Section: Figurementioning

confidence: 99%

Section: Robotics and Automation Of Crystallization Setups And Their mentioning

confidence: 99%

Introduction to protein crystallization

McPherson

Gavira

2013

Acta Crystallogr F Struct Biol Commun

357

323

View full text Add to dashboard Cite

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“…those conditions with B values within the crystallization slot). The approach of using an incomplete factorial (Kendrick et al, 1997) of crystallization conditions and outcomes combined with ANN analysis has previously been reported by DeLucas and coworkers (DeLucas et al, 2003(DeLucas et al, , 2005. The ability to improve or maximize the solubility of purified proteins is beneficial to many different protein research disciplines.…”

Section: Figurementioning

confidence: 99%

“…By combining ANN analysis of an incomplete factorial screen that surveys multiple parameter combinations and concentrations, the total number of screening conditions can be dramatically reduced. The use of an ANN and incomplete factorial screens have proven to be effective in crystallization, although the predicted novel crystallization conditions typically resulted in a large number of false positives (DeLucas et al, 2003(DeLucas et al, , 2005. However, the application of ANN and incomplete factorial solubility screens has proven to be more promising.…”

Section: Figurementioning

confidence: 99%

Applications of the second virial coefficient: protein crystallization and solubility

Wilson

DeLucas

2014

Acta Crystallogr F Struct Biol Commun

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This article begins by highlighting some of the ground-based studies emanating from NASA's Microgravity Protein Crystal Growth (PCG) program. This is followed by a more detailed discussion of the history of and the progress made in one of the NASA-funded PCG investigations involving the use of measured second virial coefficients (Bvalues) as a diagnostic indicator of solution conditions conducive to protein crystallization. A second application of measuredBvalues involves the determination of solution conditions that improve or maximize the solubility of aqueous and membrane proteins. These two important applications have led to several technological improvements that simplify the experimental expertise required, enable the measurement of membrane proteins and improve the diagnostic capability and measurement throughput.

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“…The large amounts of sample required make systematic exploration by conventional techniques infeasible, and screening is typically directed toward an incomplete factorial or sparse-matrix approach, a brute-force process requiring large numbers of experiments (15, 16). There have been numerous attempts to rationalize this procedure, for example, by using computational approaches to predict phase behavior (17,18) or by trying to correlate measurements of osmotic second virial coefficients (19, 20) with crystallization conditions. Practical limitations have thus far prevented these techniques from being generally applicable.…”

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confidence: 99%

Systematic investigation of protein phase behavior with a microfluidic formulator

Hansen

Sommer

Quake

2004

Proc. Natl. Acad. Sci. U.S.A.

178

179

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We demonstrated a microfluidic device for rapidly generating complex mixtures of 32 stock reagents in a 5-nl reactor. This ''formulation chip'' is fully automated and allows thousands of experiments to be performed in a single day with minimal reagent consumption. It was applied to systematically study the phase behavior of the protein xylanase over a large and complex chemical space. For each chemical formulation that demonstrated a pronounced effect on solubility, the protein phase behavior was completely mapped in the chip, generating a set of empirical phase diagrams. This ab initio phase information was used to devise a rational crystallization screen that resulted in 72-fold improvement in successful crystallization hits compared with conventional sparse matrix screens. This formulations tool allows a physicsbased approach to protein crystallization that may prove useful in structural genomics efforts.T he application of x-ray crystallography to the determination of protein structure with atomic resolution was a triumph of structural biology in the 20th century. Since the first solution of the structure of myoglobin in 1958 (1), Ͼ23,000 different structures have been deposited in the Protein Data Bank, and their role in relating structure to function in biology has been profound. As structure determination efforts continue to move past the most tractable crystallization targets (typically small soluble proteins) and focus instead on more challenging macromolecules, such as large protein complexes and membrane proteins (2), the need to better understand and explore the crystallization process has become urgent. That is because once high-quality crystals are in hand, advances in x-ray sources, computer codes, and related technology have made it relatively straightforward to obtain the structure. Determining the appropriate crystallization conditions has become one of the most significant remaining bottlenecks to structure determination (3).Understanding the phase behavior of proteins is an essential part of the crystallization process. The growth of crystals from a protein solution requires the existence of a nontrivial phase diagram, which allows the protein state to be manipulated between at least two thermodynamic phases: soluble and precipitated. The processes of crystal nucleation and growth arise on the boundary between these two phases and are governed by subtle effects in physical chemistry. There are a variety of schemes that manipulate the kinetics of the crystallization process, and all take advantage of generic features of these phase diagrams (4). However, in practice, the phase behavior of very few proteins has been studied in detail (5-12), and solubility information for a specific protein is rarely available for crystallization and optimization experiments (13,14).Furthermore, it is often an arduous process to find the right combination of chemicals that yields appropriate phase behavior for a given protein. Every protein is different, and even a modest subset of stock precipitating solutions...

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Efficient protein crystallization

Cited by 71 publications

References 19 publications

Introduction to protein crystallization

Introduction to protein crystallization

Applications of the second virial coefficient: protein crystallization and solubility

Systematic investigation of protein phase behavior with a microfluidic formulator

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