Abstract
Large, high-quality protein crystals are required for the structural determination of proteins via X-ray diffraction. In this article, we propose a technique to facilitate the production of such crystals and validate its feasibility through simulations. An analytical method for protein aqueous solution based on a Fourier transform infrared (FTIR) spectroscopy is combined with a temperature control strategy to manipulate the extent of supersaturation during crystal growth, thus impacting crystal quality. The technique requires minimal knowledge about the growth kinetics a priori. The simulations show that, under ideal conditions, the design can be as effective as predesigned temperature programs for crystallization based on known growth kinetics. Two kinds of errors might be encountered with this design. Error in the estimated number of seed crystals can result in a growth rate deviating from the desired one. Nevertheless, the deviation is usually tolerable and system instability is unlikely to occur. Based on the standard error of our FTIR method, errors in concentration measurement are simulated. Measurement error can result in system instability and impair the control algorithm. Such errors may be compensated by limiting the temperature change taken by the control action, or by improving the measurement precision through the use of regressed concentrations. Through simulations, it is shown that the proposed design is practical and represents a significant improvement over the commonly used isothermal crystallization technique.
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Abbreviations
- Ā :
-
average peak area of all calibration sample spectra
- A :
-
integrated peak area of protein FTIR spectra
- b :
-
empirical constant in van’t Hoff equation (1/K)
- b 0, b 1 :
-
coefficient of linear relation between A and C
- C :
-
soluble protein concentration (mg/mL)
- C*:
-
solubility of protein (mg/mL)
- G :
-
growth rate of a crystal (µm/h)
- k :
-
proportional constant for crystal growth kinetics ([µm/min]/[mg/mL])n
- k v :
-
shape factor of crystals
- L :
-
length of a crystal (µm)
- M :
-
number of samples used in concentration calibration in FTIR method
- m :
-
overall mass of crystals (mg)
- N :
-
number of crystals
- N′:
-
estimation of number of crystals
- n :
-
power law constant for crystal growth kinetics
- R :
-
ideal gas constant (0.001986 kcal/[mol·K])
- S AA :
-
sum of square of peak area from all calibration sample spectra
- T :
-
temperature (K)
- t :
-
time (h)
- t α/2 :
-
student’s t distribution
- V :
-
liquid phase volume (mL)
- ΔH cryst :
-
enthalpy of formation of crystals (kcal/mol)
- ρ eff :
-
density of crystals (mg of protein/µm3 of crystal volume)
- σ:
-
SEC in FTIR method
- ini :
-
initial conditions
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Hu, SY.B., Wiencek, J.M. & Arnold, M.A. Application of near-infrared spectra on temperature-controlled protein crystallization. Appl Biochem Biotechnol 94, 179–196 (2001). https://doi.org/10.1385/ABAB:94:2:179
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DOI: https://doi.org/10.1385/ABAB:94:2:179