Trypsin modification by vinyl polymers with variable solubilities in response to external signals

Ito, Yoshihiro; Kotoura, Masaaki; Chung, Dong June; Imanishi, Yukio

doi:10.1021/bc00023a009

Cited by 26 publications

(11 citation statements)

References 12 publications

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“…16 When the conjugates are dissolved, they efficiently catalyze reactions in the homogeneous phase where a high activity is usually achieved, and at any time they can be easily separated from the solution by inducing phase separation, e.g., by changing temperature 17 or pH. [18][19][20][21] When the enzyme is conjugated to reactive polymer sites along the polymer backbone, the active site may be partially blocked owing to steric hindrance by the polymers, with resultant loss in enzyme activity. Therefore, in the work we describe here, we conjugated a thermally sensitive oligomer by one reactive end group only to the enzyme.…”

Section: Introductionmentioning

confidence: 99%

Unusual properties of thermally sensitive oligomer-enzyme conjugates of poly(N-isopropylacrylamide)-trypsin

Ding¹,

Chen²,

Hoffman³

1998

J. Biomed. Mater. Res.

104

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Reversible soluble-insoluble oligomer-enzyme conjugates have been prepared by conjugating a thermally sensitive oligomer, poly(N-isopropylacrylamide) [poly(NIPAAm)] to trypsin. The conjugates can catalyze enzymatic reactions in solution and then may be separated from the solution by thermal precipitation. One special feature of the conjugates is that every poly(NIPAAm) chain has only one end attachment to the enzyme, so that the loss of enzymatic activity due to steric hindrance should be minimized. Conjugates with various numbers of oligomer chains per trypsin molecule were prepared. Surprisingly, the conjugates increased in enzymatic activity with increasing oligomer conjugation to the native trypsin. The trypsin active sites in the conjugates were accessible to large molecules, such as soybean trypsin inhibitor (MW = 21,500). The enzyme conjugates were more stable than native trypsin, both in solution and in the precipitated phase. On the other hand, the conjugates lost enzymatic activity faster than native trypsin when the temperature was repeatedly cycled through the lower critical solution temperature (LCST) of the poly(NIPAAm). The recovery of the conjugates by thermal precipitation in each cycle was over 95% even after 14 cycles through the LCST.

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Section: Introductionmentioning

confidence: 99%

Unusual properties of thermally sensitive oligomer-enzyme conjugates of poly(N-isopropylacrylamide)-trypsin

Ding¹,

Chen²,

Hoffman³

1998

J. Biomed. Mater. Res.

104

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“…Only a few reports have involved its application in a grafting-from method. More specifically, azo initiators immobilized on trypsin have been used to polymerize redox-or pH-sensitive monomers by radical polymerization 106 . More recently, tert-butyl hydroperoxide was used as an initiator to polymerize NIPAM on casein, and temperature and pH dual-responsive polymer-protein conjugates were prepared by polymerizing hydroxyethyl methacrylate and acrylic acid in the presence of BSA under gamma radiation 107,108 .…”

Section: Synthetic Strategies For Bioconjugationmentioning

confidence: 99%

Smart hybrid materials by conjugation of responsive polymers to biomacromolecules

et al. 2014

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The chemical structure and function of biomacromolecules has evolved to fill many essential roles in biological systems. More specifically, proteins, peptides, nucleic acids and polysaccharides serve as vital structural components, and mediate chemical transformations and energy/information storage processes required to sustain life. In many cases, the properties and applications of biological macromolecules can be further expanded by attaching synthetic macromolecules. The modification of biomacromolecules by attaching a polymer that changes its properties in response to environmental variations, thus affecting the properties of the biomacromolecule, has led to the emergence of a new family of polymeric biomaterials. Here, we summarize techniques for conjugating responsive polymers to biomacromolecules and highlight applications of these bioconjugates reported so far. In doing so, we aim to show how advances in synthetic tools could lead to rapid expansion in the variety and uses of responsive bioconjugates.

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“…Although some methods of chemical modification for regulating enzyme solubility in aqueous solution have been reported (these include the attachment of pH- 11,12 , ionic strength-13 , temperature- [14][15][16] , or redox-sensitive 12 polymers), until now there have been no reports of modifications to allow such regulation in organic media.…”

Section: Researchmentioning

confidence: 99%

Enzyme modification by polymers with solublities that change in response to photoirradiation in organic media

et al. 1999

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We have synthesized a hybrid subtilisin the solubility of which can be regulated by photoirradiation through coupling with a photoresponsive copolymer that carries spiropyran groups in its side chains. The copolymer was synthesized by polymerization of methacrylate, methacrylic acid, and spiropyran-carrying methacrylate. It was then covalently bonded to the amino groups of subtilisin Carlsberg via its carboxyl groups using a carbodiimide coupling agent. The hybrid subtilisin was perfectly soluble in toluene and efficiently catalyzed transesterification. After ultraviolet irradiation, the hybrid subtilisin precipitated and was easily and quantitatively recovered by centrifugation. Recovered hybrid subtilisin, resolubilized by visible light irradiation, retained its original transesterification activity even after several cycles of precipitation and solubilization.

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Trypsin modification by vinyl polymers with variable solubilities in response to external signals

Cited by 26 publications

References 12 publications

Unusual properties of thermally sensitive oligomer-enzyme conjugates of poly(N-isopropylacrylamide)-trypsin

Unusual properties of thermally sensitive oligomer-enzyme conjugates of poly(N-isopropylacrylamide)-trypsin

Smart hybrid materials by conjugation of responsive polymers to biomacromolecules

Enzyme modification by polymers with solublities that change in response to photoirradiation in organic media

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