Linked Life Data

18092457

Source:http://linkedlifedata.com/resource/pubmed/id/18092457

Statements in which the resource exists as a subject.
PredicateObject
rdf:type
lifeskim:mentions
pubmed:issue
11
pubmed:dateCreated
2007-12-20
pubmed:abstractText
Elastin-like polypeptides (ELPs) undergo a reversible inverse phase transition upon a change in temperature. This thermally triggered phase transition allows for a simple and rapid means of purifying a fusion protein. Recovery of ELPs-tagged fusion protein was easily achieved by aggregation, triggered either by raising temperature or by adding salt. In this study, levansucrase has been used as a model enzyme in the development of a simple one-step purification method using ELPs. The levansucrase gene cloned from Pseudomonas aurantiaca S-4380 was tagged with various sizes of ELPs to functionally express and optimize the purification of levansucrase. One of two ELPs, ELP[V-20] or ELP[V-40], was fused at the C-terminus of the levansucrase gene. A levansucrase-ELP fusion protein was expressed in Escherichia coli DH5alpha at 37 degrees C for 18 h. The molecular masses of levansucrase-ELP[V-20] and levansucrase-ELP[V-40] were determined as 56 kDa and 65 kDa, respectively. The phase transition of levansucrase-ELP[V-20] occurred at 20 degrees C in 50 mM Tris-Cl (pH 8) buffer with 3 M NaCl added, whereas the phase transition temperature (Tt) of levansucrase-ELP[V-40] was 17 degrees C with 2 M NaCl. Levansucrase was successfully purified using the phase transition characteristics of ELPs, with a recovery yield of higher than 80%, as verified by SDS-PAGE. The specific activity was measured spectrophotometrically to be 173 U/mg and 171 U/mg for levansucrase-ELP[V-20] and levansucrase-ELP[V-40], respectively, implying that the ELP-tagging system provides an efficient one-step separation method for protein purification.
pubmed:language
eng
pubmed:journal
pubmed:citationSubset
IM
pubmed:chemical
pubmed:status
MEDLINE
pubmed:month
Nov
pubmed:issn
1017-7825
pubmed:author
pubmed:issnType
Print
pubmed:volume
17
pubmed:owner
NLM
pubmed:authorsComplete
Y
pubmed:pagination
1751-7
pubmed:meshHeading
pubmed:year
2007
pubmed:articleTitle
Heterologous expression and optimized one-step separation of levansucrase via elastin-like polypeptides tagging system.
pubmed:affiliation
Department of Biological Engineering, Inha University, Incheon 402-751, Korea.
pubmed:publicationType
Journal Article