Optimization of protein solution by a novel experimental design method using thermodynamic properties.
In this study, the structural stability of hen egg white lysozyme in solution at various pH levels and in different types of buffers, including acetate, phosphate, histidine, and Tris, was investigated by means of differential scanning calorimetry (DSC). Reasonable pH values were selected from the buffer ranges and were analyzed statistically through design of experiment (DoE). Four factors were used to characterize the thermograms: calorimetric enthalpy (ΔH), temperature at maximum heat flux (T( m )), van't Hoff enthalpy (ΔH( V )), and apparent activation energy of protein solution (E(app)). It was possible to calculate E(app) through mathematical elaboration from the Lumry-Eyring model by changing the scan rate. The transition temperature of protein solution, T( m ), increased when the scan rate was faster. When comparing the T( m ), ΔH( V ), ΔH, and E(app) of lysozyme in various pH ranges and buffers with different priorities, lysozyme in acetate buffer at pH 4.767 (scenario 9) to pH 4.969 (scenario 11) exhibited the highest thermodynamic stability. Through this experiment, we found a significant difference in the thermal stability of lysozyme in various pH ranges and buffers and also a new approach to investigate the physical stability of protein by DoE.
College of Pharmacy, Pusan National University, Busan, 609-735, Korea.
Nam Ah
NA
Kim
In Bok
IB
An
Sang Yeol
SY
Lee
Eun-Seok
ES
Park
Seong Hoon
SH
Jeong
D019342:Acetic Acid / Q000737:chemistry; D000818:Animals; D030161:Avian Proteins / Q000737:chemistry*; D002021:Buffers; D002152:Calorimetry, Differential Scanning; D002626:Chemistry, Pharmaceutical; D002645:Chickens; D004355:Drug Stability; D004795:Enzyme Stability; D006358:Hot Temperature; D006863:Hydrogen-Ion Concentration; D008958:Models, Molecular; D008970:Molecular Weight; D009113:Muramidase / Q000737:chemistry*; D017510:Protein Folding; D017434:Protein Structure, Tertiary; D058767:Protein Unfolding; D019346:Sodium Acetate / Q000737:chemistry; D012996:Solutions; D013816:Thermodynamics
D030161:Avian Proteins; D002021:Buffers; D012996:Solutions; D019346:Sodium Acetate; C421864:hen egg lysozyme; D009113:Muramidase; D019342:Acetic Acid