Crystallization Experiments (crystallization + experiment)

Distribution by Scientific Domains


Selected Abstracts


Kinetic studies on the influence of temperature and growth rate history on crystal growth

CRYSTAL RESEARCH AND TECHNOLOGY, Issue 12 2008
P. M. Martins
Abstract Crystallization experiments of sucrose were performed in a batch crystallizer to study the effect of temperature and growth rate history on the crystal growth kinetics. In one of the growth methods adopted, the isothermal volumetric growth rate (RV) is determined as a function of supersaturation (S) at 35, 40 and 45 C. In the other, crystals are allowed to grow at constant supersaturation by automatically controlling the solution temperature as the solute concentration decreased. Using the latter method RV is calculated as the solution is cooled. The obtained results are interpreted using empirical, engineering and fundamental perspectives of crystal growth. Firstly, the overall activation energy (EA) is determined from the empirical growth constants obtained in the isothermal method. The concept of falsified kinetics, widely used in chemical reaction engineering, is then extended to the crystal growth of sucrose in order to estimate the true activation energy (ET) from the diffusion-affected constant, EA. The differences found in the isothermal and constant supersaturation methods are explained from the viewpoint of the spiral nucleation mechanism, taking into account different crystal surface properties caused by the growth rate history in each method. Finally, the crystal growth curve obtained in the batch crystallizer at 40 C is compared with the one obtained in a fluidized bed crystallizer at the same temperature. Apparently divergent results are explained by the effects of crystal size, hydrodynamic conditions and growth rate history on the crystallization kinetics of sucrose. ( 2008 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim) [source]


Growth of large protein crystals by a large-scale hanging-drop method

JOURNAL OF APPLIED CRYSTALLOGRAPHY, Issue 4 2010
Keisuke Kakinouchi
A method for growing large protein crystals is described. In this method, a cut pipette tip is used to hang large-scale droplets (maximum volume 200,l) consisting of protein and precipitating agents. A crystal grows at the vapor,liquid interface; thereafter the grown crystal can be retrieved by droplet,droplet contact both for repeated macroseeding and for mounting crystals in a capillary. Crystallization experiments with peroxiredoxin of Aeropyrum pernix K1 (thioredoxin peroxidase, ApTPx) and hen egg white lysozyme demonstrated that this large-scale hanging-drop method could produce a large-volume crystal very effectively. A neutron diffraction experiment confirmed that an ApTPx crystal (6.2,mm3) obtained by this method diffracted to beyond 3.5, resolution. [source]


A simple technique to convert sitting-drop vapor diffusion into hanging-drop vapor diffusion by solidifying the reservoir solution with agarose

JOURNAL OF APPLIED CRYSTALLOGRAPHY, Issue 5 2009
Tae Woong Whon
A simple protocol to convert sitting-drop vapor-diffusion plating into a hanging-drop vapor-diffusion experiment in protein crystallization is reported. After making a sitting-drop plate, agarose solution was added to solidify the reservoir solution, and the plates were incubated upside down. Crystallization experiments with hen egg white lysozyme, thaumatin and glucose isomerase showed that the `upside-down sitting-drop' method could produce single crystals with all the benefits of the hanging-drop crystallization method. [source]


Molecular Modeling in Crystal Engineering for Processing of Energetic Materials

PROPELLANTS, EXPLOSIVES, PYROTECHNICS, Issue 6 2003
Stphane Bnazet
Abstract Nowadays molecular modeling is available to explain molecular phenomena. This approach helps to compute crystal surface property effects that can be used both for morphology studies and optimal design of "bonding agents" to prevent filler-binder detachment. The principles of crystal growth and of interaction energy computing have been applied to Hexanitrohexaazaisowurtzitane (HNIW). Crystallization experiments validate our calculations. Three families of additives of crystal growth are distinguished: the retarding (and inhibitor) agents, the promoters and finally the "tailor-mades". Retarding and inhibitor agents are the most interesting one to find bonding agents. HNIW is used to present our methodology, but engineering using molecular modeling could be generalized to other fillers. [source]


Crystallization and preliminary X-ray diffraction analysis of the dimerization domain of the tumour suppressor ING4

ACTA CRYSTALLOGRAPHICA SECTION F (ELECTRONIC), Issue 5 2010
Simone Culurgioni
Inhibitor of growth protein 4 (ING4) belongs to the ING family of tumour suppressors and is involved in chromatin remodelling, in growth arrest and, in cooperation with p53, in senescence and apoptosis. Whereas the structure and histone H3-binding properties of the C-terminal PHD domains of the ING proteins are known, no structural information is available for the N-terminal domains. This domain contains a putative oligomerization site rich in helical structure in the ING2,5 members of the family. The N-terminal domain of ING4 was overexpressed in Escherichia coli and purified to homogeneity. Crystallization experiments yielded crystals that were suitable for high-resolution X-ray diffraction analysis. The crystals belonged to the orthorhombic space group C222, with unit-cell parameters a = 129.7, b = 188.3, c = 62.7,. The self-rotation function and the Matthews coefficient suggested the presence of three protein dimers per asymmetric unit. The crystals diffracted to a resolution of 2.3, using synchrotron radiation at the Swiss Light Source (SLS) and the European Synchrotron Radiation Facility (ESRF). [source]


Enzyme-assisted physicochemical enantioseparation processes,Part III: Overcoming yield limitations by dynamic kinetic resolution of asparagine via preferential crystallization and enzymatic racemization

BIOTECHNOLOGY & BIOENGINEERING, Issue 6 2009
Kerstin Wrges
Abstract The application of enantioseparation methods alone can only yield up to 50% of the desired chiral product. Thus enantioseparation becomes more attractive when accompanied by the racemization of the counter-enantiomer. Here we present first results of dynamic kinetic resolution of L -asparagine (L -Asn) via preferential crystallization and enzymatic racemization from a racemic, supersaturated solution on a 20,mL scale. An enzyme lyophilisate (WT amino acid racemase from P. putida KT2440 (E.C. 5.1.1.10), overexpressed in E. coli BL21(DE3)) was used for in situ racemization (enzyme concentrations varying from 0 to 1,mg/mL). When preferential crystallization was applied without any enzyme, a total of 31,mg of L -Asn monohydrate could be crystallized, before crystal formation of d -Asn started. Crystallization experiments accompanied by enzymatic racemization led to a significant increase of crystallized L -Asn (198,mg L -Asn monohydrate; >92%ee) giving the first experimental proof for this new process concept of dynamic kinetic resolution via preferential crystallization and enzymatic racemization. Measurements of the racemase activity before and after the crystallization process showed no significant differences, which would allow for enzyme recovery and recycling. Biotechnol. Bioeng. 2009; 104: 1235,1239. 2009 Wiley Periodicals, Inc. [source]


Autolabo: an automated system for ligand-soaking experiments with protein crystals

JOURNAL OF APPLIED CRYSTALLOGRAPHY, Issue 4 2010
Michihiro Sugahara
Ligand soaking of protein crystals is important for the preparation of heavy-atom derivative crystals for experimental phasing as well as for large-scale ligand screening in pharmaceutical developments. To facilitate laborious large-scale ligand screening, to reduce the risk of human contact with hazardous ligand reagents and to increase the success rate of the soaking experiments, a protein crystallization robot `Autolabo' has been developed and implemented in the high-throughput crystallization-to-structure pipeline at RIKEN SPring-8 Center. The main functions of this robotic system are the production of protein crystals for experiments, the ligand soaking of these crystals and the observation of soaked crystals. The separate eight-channel dispensers of Autolabo eliminate the cross-contamination of reagents which should be strictly avoided in the ligand-soaking experiment. Furthermore, the automated approach reduces physical damage to crystals during experiments when compared with the conventional manual approach, and thereby has the potential to yield better quality diffraction data. Autolabo's performance as a ligand-soaking system was evaluated with a crystallization experiment on ten proteins from different sources and a heavy-atom derivatization experiment on three proteins using a versatile cryoprotectant containing heavy-atom reagents as ligands. The crystallization test confirmed reliable crystal reproduction in a single condition and the capability for crystallization with nucleants to improve crystal quality. Finally, Autolabo reproducibly derivatized the test protein crystals with sufficient diffraction quality for experimental phasing and model building, indicating a high potentiality of this automated approach in ligand-soaking experiments. [source]


Effects of impurities on membrane-protein crystallization in different systems

ACTA CRYSTALLOGRAPHICA SECTION D, Issue 10 2009
Christopher A. Kors
When starting a protein-crystallization project, scientists are faced with several unknowns. Amongst them are these questions: (i) is the purity of the starting material sufficient? and (ii) which type of crystallization experiment is the most promising to conduct? The difficulty in purifying active membrane-protein samples for crystallization trials and the high costs associated with producing such samples require an extremely pragmatic approach. Additionally, practical guidelines are needed to increase the efficiency of membrane-protein crystallization. In order to address these conundrums, the effects of commonly encountered impurities on various membrane-protein crystallization regimes have been investigated and it was found that the lipidic cubic phase (LCP) based crystallization methodology is more robust than crystallization in detergent environments using vapor diffusion or microbatch approaches in its ability to tolerate contamination in the forms of protein, lipid or other general membrane components. LCP-based crystallizations produced crystals of the photosynthetic reaction center (RC) of Rhodobacter sphaeroides from samples with substantial levels of residual impurities. Crystals were obtained with protein contamination levels of up to 50% and the addition of lipid material and membrane fragments to pure samples of RC had little effect on the number or on the quality of crystals obtained in LCP-based crystallization screens. If generally applicable, this tolerance for impurities may avoid the need for samples of ultrahigh purity when undertaking initial crystallization screening trials to determine preliminary crystallization conditions that can be optimized for a given target protein. [source]


The plug-based nanovolume Microcapillary Protein Crystallization System (MPCS)

ACTA CRYSTALLOGRAPHICA SECTION D, Issue 11 2008
Cory J. Gerdts
The Microcapillary Protein Crystallization System (MPCS) embodies a new semi-automated plug-based crystallization technology which enables nanolitre-volume screening of crystallization conditions in a plasticware format that allows crystals to be easily removed for traditional cryoprotection and X-ray diffraction data collection. Protein crystals grown in these plastic devices can be directly subjected to in situ X-ray diffraction studies. The MPCS integrates the formulation of crystallization cocktails with the preparation of the crystallization experiments. Within microfluidic Teflon tubing or the microfluidic circuitry of a plastic CrystalCard, ,10,20,nl volume droplets are generated, each representing a microbatch-style crystallization experiment with a different chemical composition. The entire protein sample is utilized in crystallization experiments. Sparse-matrix screening and chemical gradient screening can be combined in one comprehensive `hybrid' crystallization trial. The technology lends itself well to optimization by high-granularity gradient screening using optimization reagents such as precipitation agents, ligands or cryoprotectants. [source]


Tapping the Protein Data Bank for crystallization information

ACTA CRYSTALLOGRAPHICA SECTION D, Issue 12 2005
Thomas S. Peat
A database application has been developed for the collection of crystallographic information. This database (the BDP) has been populated with the information found in the Protein Data Bank (PDB). The tool has been used to store crystallization data parsed out of the PDB and these data may be used to extend the crystallization information found in the Biological Macromolecule Crystallization Database (BMCD) and could be used to refine crystallization methodology. A standard is proposed for describing a crystallization experiment that will ease future crystallization data collations and analyses. [source]


Efficient UV detection of protein crystals enabled by fluorescence excitation at wavelengths longer than 300,nm

ACTA CRYSTALLOGRAPHICA SECTION F (ELECTRONIC), Issue 4 2010
Karsten Dierks
It is well known that most proteins and many other biomolecules fluoresce when illuminated with UV radiation, but it is also commonly accepted that utilizing this property to detect protein crystals in crystallization setups is limited by the opacity of the materials used to contain and seal them. For proteins, this fluorescence property arises primarily from the presence of tryptophan residues in the sequence. Studies of protein crystallization results in a variety of setup configurations show that the opacity of the containment hardware can be overcome at longer excitation wavelengths, where typical hardware materials are more transparent in the UV, by the use of a powerful UV-light source that is effective in excitation even though not at the maximum of the excitation response. The results show that under these circumstances UV evaluation of crystallization trials and detection of biomolecular crystals in them is not limited by the hardware used. It is similarly true that a deficiency in tryptophan or another fluorescent component that limits the use of UV light for these purposes can be effectively overcome by the addition of fluorescent prostheses that bind to the biomolecule under study. The measurements for these studies were made with a device consisting of a potent UV-light source and a detection system specially adapted (i) to be tunable via a motorized and software-controlled absorption-filter system and (ii) to convey the excitation light to the droplet or capillary hosting the crystallization experiment by quartz-fibre light guides. [source]


MyCrystals, a simple visual data management program for laboratory-scale crystallization experiments

JOURNAL OF APPLIED CRYSTALLOGRAPHY, Issue 4 2009
Monika Nhr Lvgreen
MyCrystals is designed as a user-friendly program to display crystal images and list crystallization conditions. The crystallization conditions entry fields can be customized to suit the experiments. MyCrystals is also able to sort the images by the entered crystallization conditions, which presents a unique opportunity to easily assess the effect of, for example, changing pH or concentration and thus establish the best conditions to be used for optimization. [source]


Automated classification of crystallization experiments using wavelets and statistical texture characterization techniques

JOURNAL OF APPLIED CRYSTALLOGRAPHY, Issue 1 2008
D. Watts
A method is presented for the classification of protein crystallization images based on image decomposition using the wavelet transform. The distribution of wavelet coefficient values in each sub-band image is modelled by a generalized Gaussian distribution to provide discriminatory variables. These statistical descriptors, together with second-order statistics obtained from joint probability distributions, are used with learning vector quantization to classify protein crystallization images. [source]


Nonlinear kinetic parameter estimation for batch cooling seeded crystallization

AICHE JOURNAL, Issue 8 2004
Q. Hu
Abstract Kinetic parameter estimation for most batch crystallization processes is necessary because nucleation and crystal growth kinetic parameters are often not available. The existing identification methods are generally based on simplified population balance models such as moment equations, which contain insufficient information on the crystal size distribution (CSD). To deal with these problems, a new optimization-based identification approach for general batch cooling seeded crystallization is proposed in this study. The final-time CSD is directly used for identification. A novel effective method for solving the population balance equation is developed and used to identify nucleation and growth kinetic parameters. Cooling crystallization of ammonium sulfate in water was experimentally investigated, where the concentration was measured by an on-line density meter and the final-time CSD was analyzed by a Malvern Mastersizer 2000. Kinetics for ammonium sulfate are determined based on cooling crystallization experiments. Applying these kinetics in simulation provides a good prediction of the product CSD. 2004 American Institute of Chemical Engineers AIChE J, 50: 1786,1794, 2004 [source]


Antisolvent crystallization of anhydrous sodium carbonate at atmospherical conditions

AICHE JOURNAL, Issue 3 2001
Harald Oosterhof
When antisolvents are applied to crystallize sodium carbonate from aqueous solutions, the transition temperature at which the hydrates are in equilibrium is decreased. Two models proposed can predict the influence of the amount and type of antisolvent on the transition temperature. Only binary data of the water/sodium carbonate system and measured vapor pressures over ternary soda-saturated mixtures of water and antisolvent are needed. To validate the two models, continuous crystallization experiments were carried out at various temperatures using ethylene glycol (EG) and diethylene glycol (DEG) as antisolvent, in varying concentrations. Both models predict the influence of the antisolvent on the transition temperature with good accuracy. Anhydrous soda with bulk densities of up to 950 kg/m3 was crystallized at temperatures as low as 80C. [source]


Mechanistic Investigation into the Unique Orientation Textures of Poly(vinylidene fluoride) in Blends with Nylon 11

MACROMOLECULAR RAPID COMMUNICATIONS, Issue 10 2003
Yongjin Li
Abstract Self-seeded crystallization experiments were carried out to detect the mechanism of the unique orientation behavior of poly(vinylidene fluoride) (PVDF) in oriented PVDF/nylon 11 blends. It was found that primary nuclei have no effects on the final orientation textures adopted by PVDF. The results show that the PVDF crystal orientation in the oriented blends is determined in the early stage of crystal growth, thus a trans crystallization mechanism is preferred. Isothermal crystallization kinetics for the self-seeded and non-self-seeded crystallization at 145,C. [source]


Quantitive evaluation of macromolecular crystallization experiments using 1,8-ANS fluorescence

ACTA CRYSTALLOGRAPHICA SECTION D, Issue 8 2010
David Watts
Modern X-ray structure analysis and advances in high-throughput robotics have allowed a significant increase in the number of conditions screened for a given sample volume. An efficient evaluation of the increased amount of crystallization trials in order to identify successful experiments is now urgently required. A novel approach is presented for the visualization of crystallization experiments using fluorescence from trace amounts of a nonspecific dye. The fluorescence images obtained strongly contrast protein crystals against other phenomena, such as precipitation and phase separation. Novel software has been developed to quantitatively evaluate the crystallization outcome based on a biophysical metric correlated with voxel protein concentration. In >1500 trials, 85.6% of the successful crystallization experiments were correctly identified, yielding a 50% reduction in the number of `missed hits' compared with current automated approaches. The use of the method in the crystallization of three previously uncharacterized proteins from the malarial parasite Plasmodium falciparum is further demonstrated. [source]


The plug-based nanovolume Microcapillary Protein Crystallization System (MPCS)

ACTA CRYSTALLOGRAPHICA SECTION D, Issue 11 2008
Cory J. Gerdts
The Microcapillary Protein Crystallization System (MPCS) embodies a new semi-automated plug-based crystallization technology which enables nanolitre-volume screening of crystallization conditions in a plasticware format that allows crystals to be easily removed for traditional cryoprotection and X-ray diffraction data collection. Protein crystals grown in these plastic devices can be directly subjected to in situ X-ray diffraction studies. The MPCS integrates the formulation of crystallization cocktails with the preparation of the crystallization experiments. Within microfluidic Teflon tubing or the microfluidic circuitry of a plastic CrystalCard, ,10,20,nl volume droplets are generated, each representing a microbatch-style crystallization experiment with a different chemical composition. The entire protein sample is utilized in crystallization experiments. Sparse-matrix screening and chemical gradient screening can be combined in one comprehensive `hybrid' crystallization trial. The technology lends itself well to optimization by high-granularity gradient screening using optimization reagents such as precipitation agents, ligands or cryoprotectants. [source]


Improved classification of crystallization images using data fusion and multiple classifiers

ACTA CRYSTALLOGRAPHICA SECTION D, Issue 8 2008
Samarasena Buchala
Identifying the conditions that will produce diffraction-quality crystals can require very many crystallization experiments. The use of robots has increased the number of experiments performed in most laboratories, while in structural genomics centres tens of thousands of experiments can be produced every day. Reliable automated evaluation of these experiments is becoming increasingly important. A more robust classification is achieved by combining different methods of feature extraction with the use of multiple classifiers. [source]


Heterogeneous nucleation of three-dimensional protein nanocrystals

ACTA CRYSTALLOGRAPHICA SECTION D, Issue 5 2007
Dilyana G. Georgieva
Nucleation is the rate-limiting step in protein crystallization. Introducing heterogeneous substrates may in some cases lower the energy barrier for nucleation and thereby facilitate crystal growth. To date, the mechanism of heterogeneous protein nucleation remains poorly understood. In this study, the nucleating properties of fragments of human hair in crystallization experiments have been investigated. The four proteins that were tested, lysozyme, glucose isomerase, a polysaccharide-specific Fab fragment and potato serine protease inhibitor, nucleated preferentially on the hair surface. Macrocrystals and showers of tiny crystals of a few hundred nanometres thickness were obtained also under conditions that did not produce crystals in the absence of the nucleating agent. Cryo-electron diffraction showed that the nanocrystals diffracted to at least 4, resolution. The mechanism of heterogeneous nucleation was studied using confocal fluorescent microscopy which demonstrated that the protein is concentrated on the nucleating surface. A substantial accumulation of protein was observed on the sharp edges of the hair's cuticles, explaining the strong nucleating activity of the surface. [source]


An automated image-collection system for crystallization experiments using SBS standard microplates

ACTA CRYSTALLOGRAPHICA SECTION D, Issue 2 2007
Erik Brostromer
As part of a structural genomics platform in a university laboratory, a low-cost in-house-developed automated imaging system for SBS microplate experiments has been designed and constructed. The imaging system can scan a microplate in 2,6,min for a 96-well plate depending on the plate layout and scanning options. A web-based crystallization database system has been developed, enabling users to follow their crystallization experiments from a web browser. As the system has been designed and built by students and crystallographers using commercially available parts, this report is aimed to serve as a do-it-yourself example for laboratory robotics. [source]


Evaluation of crystalline objects in crystallizing protein droplets based on line-segment information in greyscale images

ACTA CRYSTALLOGRAPHICA SECTION D, Issue 3 2006
Kuniaki Kawabata
Several automated crystallization systems have recently been developed for high-throughput X-ray structure analysis. However, the evaluation process for the growth state of crystallizing protein droplets has not yet been completely automated. This paper proposes a new evaluation method for crystalline objects in automated crystallization experiments. The main objective is to determine whether a droplet contains crystals suitable for diffraction experiments and analysis. The evaluation method developed here involves extracting line-segment features from an image of the droplet and discriminating the state of crystallization using classifiers based on line features. In order to verify the efficacy of the proposed method, it was used to classify images obtained by an automated crystallization system. [source]


A procedure for setting up high-throughput nanolitre crystallization experiments.

ACTA CRYSTALLOGRAPHICA SECTION D, Issue 6 2005
Crystallization workflow for initial screening, automated storage, imaging, optimization
Crystallization trials at the Division of Structural Biology in Oxford are now almost exclusively carried out using a high-throughput workflow implemented in the Oxford Protein Production Facility. Initial crystallization screening is based on nanolitre-scale sitting-drop vapour-diffusion experiments (typically 100,nl of protein plus 100,nl of reservoir solution per droplet) which use standard crystallization screening kits and 96-well crystallization plates. For 294,K crystallization trials the barcoded crystallization plates are entered into an automated storage system with a fully integrated imaging system. These plates are imaged in accordance with a pre-programmed schedule and the resulting digital data for each droplet are harvested into a laboratory information-management system (LIMS), scored by crystal recognition software and displayed for user analysis via a web-based interface. Currently, storage for trials at 277,K is not automated and for imaging the crystallization plates are fed by hand into an imaging system from which the data enter the LIMS. The workflow includes two procedures for nanolitre-scale optimization of crystallization conditions: (i) a protocol for variation of pH, reservoir dilution and protein:reservoir ratio and (ii) an additive screen. Experience based on 592 crystallization projects is reported. [source]


An improved protocol for rapid freezing of protein samples for long-term storage

ACTA CRYSTALLOGRAPHICA SECTION D, Issue 1 2004
Rapid freezing of protein samples
Freezing of purified protein drops directly in liquid nitrogen is a convenient technique for the long-term storage of protein samples. Although this enhances reproducibility in follow-up crystallization experiments, some protein samples are not amenable to this technique. It has been discovered that plunging PCR tubes containing protein samples into liquid nitrogen results in more rapid freezing of the samples and can safely preserve some proteins that are damaged by drop-freezing. The PCR-tube method can also be adapted to a PCR-plate freezing method with applications for high-throughput and structural genomics projects. [source]


Compatibility of detergents with the microbatch-under-oil crystallization method

ACTA CRYSTALLOGRAPHICA SECTION D, Issue 6 2003
Patrick J. Loll
Detergents are required to solubilize integral membrane proteins and are common components of the solutions used to crystallize these molecules. It has been unclear whether these detergents are completely compatible with the oils used in the microbatch-under-oil crystallization technique, because they might conceivably be lost from solution by partitioning into the oil phase. The partitioning of the detergents n -octyl-,- d -glucopyranoside and Fos-Choline-12 into two different oils used for microbatch crystallization experiments has been examined. It was found that vigorous mixing and prolonged incubation of the aqueous detergent solutions with the oils leads to small losses of detergent (approximately 5% of the total detergent mass); however, gentle mixing that is more typical of the mixing encountered in a crystallization experiments leads to negligible loss of detergent. [source]


An improved strategy for the crystallization of Leishmania mexicana pyruvate kinase

ACTA CRYSTALLOGRAPHICA SECTION F (ELECTRONIC), Issue 3 2010
Hugh P. Morgan
The inclusion of novel small molecules in crystallization experiments has provided very encouraging results and this method is now emerging as a promising alternative strategy for crystallizing `problematic' biological macromolecules. These small molecules have the ability to promote lattice formation through stabilizing intermolecular interactions in protein crystals. Here, the use of 1,3,6,8-pyrenetetrasulfonic acid (PTS), which provides a helpful intermolecular bridge between Leishmania mexicana PYK (LmPYK) macromolecules in the crystal, is reported, resulting in the rapid formation of a more stable crystal lattice at neutral pH and greatly improved X-ray diffraction results. The refined structure of the LmPYK,PTS complex revealed the negatively charged PTS molecule to be stacked between positively charged (surface-exposed) arginine side chains from neighbouring LmPYK molecules in the crystal lattice. [source]


Precipitants and additives for membrane crystallization of lysozyme

BIOTECHNOLOGY JOURNAL, Issue 11 2006
Xinmiao Zhang
Abstract Membrane crystallization is a newly developed crystallization technique that has proven to be superior in producing good crystal forms under operating conditions that are not appropriate to perform the crystallization process by other traditional techniques. In this work, static membrane crystallization was carried out on lysozyme, with hollow-fiber microporous hydrophobic membranes. Numerous precipitant and additive types and concentrations were employed in the crystallization processes in order to select the most appropriate precipitant and additive types and to find their corresponding concentration levels that can yield the best crystal forms. The crystallization processes were analyzed in two ways: firstly, by evaluation of the transmembrane fluxes obtained by using different precipitants and additives; secondly, by utilization of the images and results obtained from the micrography and IR spectra in comparisons and evaluations of the crystals formed under all kinds of conditions. Moreover, the size distributions of the crystals yielded under several typical crystallization conditions were analyzed, and turbidity and induction time periods obtained during typical crystallization experiments were also measured. Amongst the numerous precipitants and additives tested, the most appropriate precipitant type and additive were chosen and their concentrations were optimized. Good lysozyme crystals were obtained using a certain precipitant and additive. The obtained results from this work further support the advantages of utilizing the membrane crystallization technique for macromolecule crystallizations. [source]