[논문]Overexpression, Purification, and Preliminary X-ray Crystallographic Analysis of Human Brain-Type Creatine Kinase

Bong, Seung-Min; Moon, Jin-Ho; Jang, Eun-Hyuk; Lee, Ki-Seog; Chi, Young-Min

Overexpression, Purification, and Preliminary X-ray Crystallographic Analysis of Human Brain-Type Creatine Kinase 원문보기

Journal of microbiology and biotechnology, v.18 no.2, 2008년, pp.295 - 298

Bong, Seung-Min (Division of Biotechnology, College of Life Sciences, Korea University) , Moon, Jin-Ho (Division of Biotechnology, College of Life Sciences, Korea University) , Jang, Eun-Hyuk (Division of Biotechnology, College of Life Sciences, Korea University) , Lee, Ki-Seog (Division of Biotechnology, College of Life Sciences, Korea University) , Chi, Young-Min (Division of Biotechnology, College of Life Sciences, Korea University)

Abstract ▼ AI-Helper

Creatine kinase (CK; E.C. 2.7.3.2) is an important enzyme that catalyzes the reversible transfer of a phosphoryl group from ATP to creatine in energy homeostasis. The brain-type cytosolic isoform of creatine kinase (BB-CK), which is found mainly in the brain and retina, is a key enzyme in brain energy metabolism, because high-energy phosphates are transfered through the creatine kinase/phosphocreatine shuttle system. The recombinant human BB-CK protein was overexpressed as a soluble form in Escherichia coli and crystallized at $22^{\circ}C$ using PEG 4000 as a precipitant. Native X-ray diffraction data were collected to $2.2{\AA}$ resolution using synchrotron radiation. The crystals belonged to the tetragonal space group $P4_32_12$, with cell parameters of a=b=97.963, $c=164.312{\AA},\;and\;{\alpha}={\beta}={\gamma}=90^{\circ}$. The asymmetric unit contained two molecules of CK, giving a crystal volume per protein mass $(V_m)$ of $1.80{\AA}^3\;Da^{-1}$ and a solvent content of 31.6%.

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제안 방법

Even though a number of crystal structures for the CK family have been solved and analyzed extensively, a definitive catalysis mechanism has not yet been elucidated: For example, the exact role of the negatively charged NEED (Asn230, Glu227, Glu232, and Asp228) motif that is important for substrate binding and catalysis; Glu232 forms a bidentate salt bridge, and is positioned to act as a base to remove a proton from the nucleophilic nitrogen [16]; and Cys283, which is related to substrate recognition and catalysis, interacts with the non-nucleophilic r]-nitrogen [14]. In order to understand isoform specific substrate binding and catalysis in the human BB-CK, we overexpressed and crystallized the human BB-CK protein, and collected Xray ciystallographic data for determination of its three dimensional structure.

대상 데이터

2 M ammonium acetate, and 30% PEG 4000, before being flash-frozen in liquid nitrogen. X-ray diffraction data were collected from the cooled crystals using an ADSC Quantum CCD 210 detector at the beamline 4A at Pohang Accelerator Labora:ory (Pohang, South Korea). Diffraction data were collected to 2.

이론/모형

The protein s_usponsion was concentrated to 40 mg/ml for crystallization. The protein concentration was determined by using Bradford's method with bovine serum albumin as the standard. The initial crystallization screening was performed using Crystal Screen I and II kits (Hampton Research, U.

참고문헌 (20)

Abraham, M. R., V. A. Selivanov, D. M. Hodgson, D. Pucar, L. V. Zingman, B. Wieringa, P. P. Dzeja, A. E. Alekseev, and A. Terzic. 2002. Coupling of cell energetics with membrane metabolic sensing. Integrative signaling through creatine kinase phosphotransfer disrupted by M-CK gene knock-out. J. Biol. Chem. 277: 24427-24434

상세보기
Aksenov, M. Y., M. V. Aksenova, R. M. Payne, C. D. Smith, W. R. Markesbery, and J. M. Carney. 1997. The expression of creatine kinase isoenzymes in neocortex of patients with neurodegenerative disorders: Alzheimer's and Pick's disease. Exp. Neurol. 146: 458-465

상세보기
Crawford, R. M., H. J. Ranki, C. H. Botting, G. R. Budas, and A. Jovanovic. 2002. Creatine kinase is physically associated with the cardiac ATP-sensitive K+ channel in vivo. FASEB J. 16: 102-104

상세보기
David, S., M. Shoemaker, and B. E. Haley. 1998. Abnormal properties of creatine kinase in Alzheimer's disease brain: Correlation of reduced enzyme activity and active site photolabeling with aberrant cytosol-membrane partitioning. Mol. Brain Res. 54: 276-287

상세보기
Eder, M., K. Fritz-Wolf, W. Kabsch, T. Wallimann, and U. Schlattner. 2000. Crystal structure of human ubiquitous mitochondrial creatine kinase. Proteins 39: 216-225

상세보기
Eder, M., U. Schlatter, A. Becker, T. Wallimann, W. Kabsch, and K. Fritz-Wolf. 1999. Crystal structure of brain-type creatine kinase at 1.41 $\AA$ resolution. Protein Sci. 8: 2258-2269

상세보기
Fritz-Wolf, K., T. Schnyder, T. Wallimann, and W. Kabsch. 1996. Structure of mitochondrial creatine kinase. Nature 381: 341-345

상세보기
Grosse, R., E. Spitzer, V. V. Kupriyanov, V. A. Saks, and K. R. Repke. 1980. Coordinate interplay between (Na+ + K+)- ATPase and creatine phosphokinase optimizes (Na+/K+)-antiport across the membrane of vesicles formed from the plasma membrane of cardiac muscle cell. Biochim. Biophys. Acta 603: 142-156

상세보기
Guerrero, M. L., J. Beron, B. Spindler, P. Groscurth, T. Wallimann, and F. Verrey. 1997. Metabolic support of Na+ pump in apically permeabilized A6 kidney cell epithelia: Role of creatine kinase. Am. J. Physiol. 272: C697-C706

상세보기
Kaldis, P., W. Hemmer, E. Zanolla, D. Holtzman, and T. Wallimann. 1996. Hot spots of creatine kinase localization in brain: Cerebellum, hippocampus and choroid plexus. Dev. Neurosci. 18: 542-554

상세보기
Lahiri, S. D., P. F. Wang, P. C. Babbitt, M. J. McLeish, G. L. Kenyon, and K. N. Allen. 2002. The 2.1 $\AA$ structure of Torpedo californica creatine kinase complexed with the $ADP-Mg^{2+}-NO^{3-}-creatine$ transition-state analogue complex. Biochemistry 41: 13861-13867

상세보기
Matthews, B. W. 1968. Solvent content of protein crystals. J. Mol. Biol. 33: 491-497

상세보기
McCoy, A. J., R. W. Grosse-Kunstleve, L. C. Storoni, and R. J. Read. 2005. Likelihood-enhanced fast translation functions. Acta Crystallogr. D 61: 458-464
Mcleish, M. J. and G. L. Kenyon. 2005. Relating structure to mechanism in creatine kinase. Biochem. Mol. Biol. 40: 1-20

상세보기
Ohren, J. F., M .L. Kundracik, C. L. Borders, P. Edmistonb, and E. R. Viola. 2007. Structural asymmetry and intersubunit communication in muscle creatine kinase. Acta Crystallogr. D 63: 381-389
Pruett, P. S., A. Azzi, S. A. Clark, M. S. Yousef, J. L. Gattis, T. Somasundaram, W. R. Ellington, and M. S. Chapman. 2003.The putative catalytic bases have, at most, an accessory role in the mechanism of arginine kinase. J. Biol. Chem. 278: 26952-26957

상세보기
Rao, J. K., G. Bujacz, and A. Wlodawer. 1998. Crystal structure of rabbit muscle creatine kinase. FEBS Lett. 439: 133-137

상세보기
Shen, Y. Q., L. Tang, H. M. Zhou, and Z. J. Lin. 2001. Structure of human muscle creatine kinase. Acta Crystallogr. D 57: 1196-1200

상세보기
Tisi, D., B. Bax, and A. Loew. 2000. The three-dimensional structure of cytosolic bovine retinal creatine kinase. Acta Crystallogr. D 57: 187-193
Walliman, T. and W. Hemmer. 1994. Creatine kinase in nonmuscle tissues and cells. Mol. Cell. Biochem. 133-134: 193-220

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