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01000naa a22002652 4500 |
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NLM175714312 |
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DE-627 |
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20231223142608.0 |
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231223s2008 xx ||||| 00| ||eng c |
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|a pubmed24n0586.xml
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|a (DE-627)NLM175714312
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|a (NLM)18076197
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|a DE-627
|b ger
|c DE-627
|e rakwb
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| 041 |
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|a eng
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| 100 |
1 |
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|a Cullen, Sean P
|e verfasserin
|4 aut
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| 245 |
1 |
0 |
|a Polymeric brushes as functional templates for immobilizing ribonuclease A
|b study of binding kinetics and activity
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| 264 |
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1 |
|c 2008
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| 336 |
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|a Text
|b txt
|2 rdacontent
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| 337 |
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|a ohne Hilfsmittel zu benutzen
|b n
|2 rdamedia
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| 338 |
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|a Band
|b nc
|2 rdacarrier
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| 500 |
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|a Date Completed 02.04.2008
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|a Date Revised 24.11.2016
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|a published: Print-Electronic
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|a Citation Status MEDLINE
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| 520 |
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|a The ability to immobilize proteins with high binding capacities on surfaces while maintaining their activity is critical for protein microarrays and other biotechnological applications. We employed poly(acrylic acid) (PAA) brushes as templates to immobilize ribonuclease A (RNase A), which is commonly used to remove RNA from plasmid DNA preparations. The brushes are grown by surface-anchored atom-transfer radical polymerization (ATRP) initiators. RNase A was immobilized by both covalent esterification and a high binding capacity metal-ion complexation method to PAA brushes. The polymer brushes immobilized 30 times more enzyme compared to self-assembled monolayers. As the thickness of the brush increases, the surface density of the RNase A increases monotonically. The immobilization was investigated by ellipsometry, X-ray photoelectron spectroscopy (XPS), thermogravimetric analysis (TGA), and near-edge X-ray absorption fine structure spectroscopy (NEXAFS). The activity of the immobilized RNase A was determined using UV absorbance. As much as 11.0 microg/cm(2) of RNase A was bound to PAA brushes by metal-ion complexation compared to 5.8 microg/cm(2) by covalent immobilization which is 30 and 16 times the estimated mass bound in a monolayer. The calculated diffusion coefficient D was 0.63 x 10(-14) cm(2)/s for metal-ion complexation and 0.71 x 10(-14) cm(2)/s for covalent immobilization. Similar values of D indicate that the binding kinetics is similar, but the thermodynamic equilibrium coverage varies with the binding chemistry. Immobilization kinetics and thermodynamics were characterized by ellipsometry for both methods. A maximum relative activity of 0.70-0.80 was reached between five and nine monolayers of the immobilized enzyme. However, the relative activity for covalent immobilization was greater than that of metal-ion complexation. Covalent esterification resulted in similar temperature dependence as free enzyme, whereas metal-ion complexation showed no temperature dependence indicating a significant change in conformation
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|a Journal Article
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| 650 |
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|a Research Support, U.S. Gov't, Non-P.H.S.
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| 650 |
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7 |
|a Acrylic Resins
|2 NLM
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| 650 |
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7 |
|a Coated Materials, Biocompatible
|2 NLM
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| 650 |
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7 |
|a Enzymes, Immobilized
|2 NLM
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| 650 |
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7 |
|a carbopol 940
|2 NLM
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| 650 |
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7 |
|a 4Q93RCW27E
|2 NLM
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| 650 |
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7 |
|a Ribonuclease, Pancreatic
|2 NLM
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| 650 |
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7 |
|a EC 3.1.27.5
|2 NLM
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| 650 |
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7 |
|a Silicon
|2 NLM
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| 650 |
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7 |
|a Z4152N8IUI
|2 NLM
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| 700 |
1 |
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|a Liu, Xiaosong
|e verfasserin
|4 aut
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| 700 |
1 |
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|a Mandel, Ian C
|e verfasserin
|4 aut
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| 700 |
1 |
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|a Himpsel, Franz J
|e verfasserin
|4 aut
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| 700 |
1 |
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|a Gopalan, Padma
|e verfasserin
|4 aut
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| 773 |
0 |
8 |
|i Enthalten in
|t Langmuir : the ACS journal of surfaces and colloids
|d 1992
|g 24(2008), 3 vom: 05. Feb., Seite 913-20
|w (DE-627)NLM098181009
|x 1520-5827
|7 nnns
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| 773 |
1 |
8 |
|g volume:24
|g year:2008
|g number:3
|g day:05
|g month:02
|g pages:913-20
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| 912 |
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|a GBV_USEFLAG_A
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| 912 |
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|a SYSFLAG_A
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| 912 |
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|a GBV_NLM
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| 912 |
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|a GBV_ILN_22
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| 912 |
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|a GBV_ILN_350
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| 912 |
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|a GBV_ILN_721
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| 951 |
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|a AR
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| 952 |
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|d 24
|j 2008
|e 3
|b 05
|c 02
|h 913-20
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