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High capacity polyelectrolyte brushe...

Kusumo, Andy T.

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High capacity polyelectrolyte brushes for tuning protein adsorption.

紀錄類型:	書目-語言資料,印刷品 : Monograph/item
正題名/作者:	High capacity polyelectrolyte brushes for tuning protein adsorption./
作者:	Kusumo, Andy T.
面頁冊數:	222 p.
附註:	Source: Dissertation Abstracts International, Volume: 69-03, Section: B, page: 1782.
Contained By:	Dissertation Abstracts International69-03B.
標題:	Biophysics, General. -
電子資源:	http://pqdd.sinica.edu.tw/twdaoapp/servlet/advanced?query=3305810
ISBN:	9780549525288

High capacity polyelectrolyte brushes for tuning protein adsorption.
Kusumo, Andy T.

High capacity polyelectrolyte brushes for tuning protein adsorption. - 222 p.

Source: Dissertation Abstracts International, Volume: 69-03, Section: B, page: 1782.

Thesis (Ph.D.)--Carnegie Mellon University, 2008.

We introduce a high capacity medium made of poly(2-(dimethylamino)ethyl methacrylate) (PDMAEMA) brushes for protein separation technology and as an enzyme immobilization matrix, and presented the physical chemistry aspects concerning its protein binding characteristics. Cationic "annealed" PDMAEMA brushes end-grafted on gold surface were able to bind a significant amount of net negatively charged model protein bovine serum albumin (BSA) at pH 6 and low ionic strength (≤ 10 mM), as indicated by surface plasmon resonance (SPR) spectroscopy measurements. BSA bound to the brush with such a high affinity that rinsing the mixed PDMAEMA/BSA layer with protein-free solution caused negligible desorption. A systematic adsorption study on PDMAEMA brushes with different degree of polymerization (Mn = 12,000--435,130) and grafting density (≤ 0.5 chains/nm2) revealed that brush binds BSA in proportion to mass of PDMAEMA, following a binding ratio of 104 DMAEMA monomers per protein, which can be considered as the "length of segment" required to wrap around one protein unit. In comparison study with the smaller size soybean trypsin inhibitor (STI), we found the "length of segment" for STI was approximately 3.7 times shorter than that of BSA. This 1:3.7 "length of segment" ratio was only partially explained by the hydrodynamic radius ratio of the corresponding proteins, which was 1:1.5 as reported from past studies. The difference in the ionic character possibly contributed to the observed 1:3 "length of segment" ratio in addition to the size difference; it might require fewer DMAEMA monomers to bind single unit of STI, which has a higher charge density than BSA.

ISBN: 9780549525288Subjects--Topical Terms:

1019105
Biophysics, General.

High capacity polyelectrolyte brushes for tuning protein adsorption.
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245 1 0 $a High capacity polyelectrolyte brushes for tuning protein adsorption.
300 $a 222 p.
500 $a Source: Dissertation Abstracts International, Volume: 69-03, Section: B, page: 1782.
502 $a Thesis (Ph.D.)--Carnegie Mellon University, 2008.
520 $a We introduce a high capacity medium made of poly(2-(dimethylamino)ethyl methacrylate) (PDMAEMA) brushes for protein separation technology and as an enzyme immobilization matrix, and presented the physical chemistry aspects concerning its protein binding characteristics. Cationic "annealed" PDMAEMA brushes end-grafted on gold surface were able to bind a significant amount of net negatively charged model protein bovine serum albumin (BSA) at pH 6 and low ionic strength (≤ 10 mM), as indicated by surface plasmon resonance (SPR) spectroscopy measurements. BSA bound to the brush with such a high affinity that rinsing the mixed PDMAEMA/BSA layer with protein-free solution caused negligible desorption. A systematic adsorption study on PDMAEMA brushes with different degree of polymerization (Mn = 12,000--435,130) and grafting density (≤ 0.5 chains/nm2) revealed that brush binds BSA in proportion to mass of PDMAEMA, following a binding ratio of 104 DMAEMA monomers per protein, which can be considered as the "length of segment" required to wrap around one protein unit. In comparison study with the smaller size soybean trypsin inhibitor (STI), we found the "length of segment" for STI was approximately 3.7 times shorter than that of BSA. This 1:3.7 "length of segment" ratio was only partially explained by the hydrodynamic radius ratio of the corresponding proteins, which was 1:1.5 as reported from past studies. The difference in the ionic character possibly contributed to the observed 1:3 "length of segment" ratio in addition to the size difference; it might require fewer DMAEMA monomers to bind single unit of STI, which has a higher charge density than BSA.
520 $a In addition to the mass of brush, tunable BSA binding capacity was demonstrated by adjusting pH and ionic strength, which manipulated electrostatic force between BSA and the brush. It was interesting that PDMAEMA was able to bind multilayers of BSA at pH 4 (ionic strengths ≤ 10 mM), or approximately one unit below its isoelectric point (pI). This "wrong side of pI" adsorption on the cationic PDMAEMA brush appeared to be weaker compared to the results reported for anionic brushes, in which adsorption occurred two pH units above isoelectric point. Recent theoretical work attributed this so-called "wrong side of pI " adsorption to (1) "charge reversal" i.e. the pH inside a cationic (anionic) brush is higher (lower) than that outside brush and (2) "counterion evaporation" i.e. binding to "patches" with opposite charges releases counterions which increases the entropy of the system.
520 $a Practical protein separation technology requires media with not only high capacity but also selectivity, regenerability, and durability. Protein selectivity based on charge was demonstrated from an adsorption study employing net positively charged lysozyme, which was completely rejected by the brush. Regeneration schemes were presented in the study. The PDMAEMA brush demonstrated good durability. At one occasion a series of 29 sequential adsorptions and washings were done on single brush without signs of capacity loss.
520 $a An attempt to create a medium with higher binding capacity was made by creating a lightly crosslinked surface-attached PDMAEMA hydrogel prepared by spin coating technique. The SPR measurements indicated it was capable of binding multilayers of BSA. The binding capacity however was not higher and the adsorption rate was lower compared to the corresponding cases measured in the brush system. Results indicated binding ratios ≥ 1000 DMAEMA monomers per protein rather than the 104 monomers per protein previously measured. The horizontal-lying segments might hinder BSA diffusion deep into the layer and/or the presence of crosslinks possibly confined the segments from conforming to the curvature of protein surface to maximize binding energy. Despite the apparent lower capacity, stronger surface attachment and simple preparation could give the hydrogel practical advantages over the brush configuration.
520 $a Temperature was not found to be a parameter one could practically use to tune protein adsorption on the PDMAEMA brush or surface-attached PDMAEMA hydrogel. Protein uptake was comparable at room temperature and at 40°C. The reason was likely that the characteristic "transition temperature" for the PDMAEMA brush was too high, as suggested by cloud point measurements. The concept of a smart surface was nevertheless demonstrated from studies which were conducted on brush system made from neutral poly(2-(2-methoxyethoxy)ethyl methacrylate) (PMEO2MA). Compared to PDMAEMA, this polymer has a more appropriate solution temperature behavior; the cloud point is near room temperature and is not pH dependent.
520 $a The impact of adsorption to the PDMAEMA brush on BSA structure was investigated by conducting solution-based studies employing fluorescence spectroscopy and circular dichroism (CD) spectroscopy. Measurements from the two spectroscopic techniques indicated BSA maintained its secondary and tertiary structure when it bound to PDMAEMA. (Abstract shortened by UMI.)
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