Crystallization of N-terminal Strep-tagged Fusion Lipase from Thermostable Bacillus sp. Strain 42.
Lipases have great potential to be used in industries due to their favourable properties such as substrate specific, enantiomerically selective, regioselective and mild reactions conditions. Lipases of microbial origin are generally more stable than lipases from animal or plant and as such they...
| Main Authors: | , , , |
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| Format: | Article |
| Language: | English |
| Published: |
IUCr
2009
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| Subjects: | |
| Online Access: | http://irep.iium.edu.my/1580/ http://irep.iium.edu.my/1580/ http://irep.iium.edu.my/1580/1/Tg_mahiran_crystal.pdf |
| Summary: | Lipases have great potential to be used in industries due
to their favourable properties such as substrate specific,
enantiomerically selective, regioselective and mild
reactions conditions. Lipases of microbial origin are
generally more stable than lipases from animal or plant and
as such they are useful source for industrial enzymes. A 1.2
kb lipase gene (AY 78735) [1], isolated from solvent stable
and thermostable Bacillus sp. strain 42 was overexpressed
using pET51b vector with E. coli host strain BL21(DE3)
pLysS, in which the fusion lipase contains N-terminal
Strep-tag II affinity tag [2]. The purified fusion lipase, at protein concentration of about 4.0 mg/mL, was induced
to crystallize in 0.1 M MES buffer at pH 6.5 without the
presence of salt, but in the presence of only 12% w/v PEG
20 000 as precipitant. Crystallization reactions were carried
out using vapour diffusion methods at 16˚C. Crystals
were formed after 12 hours incubation. The crystals with
size measuring around 0.04 X 0.12 mm were shown to be
heavily stained with protein dyes. Lip 42 lipase is highly
homologous to three crystallized lipases from thermophilic
Bacillus sp., namely T1 lipase [3], P1 lipase [4] and L1
lipase [5]. Lip 42 protein crystals, despite having almost
97% similar homology in amino acid sequence, showed
a different shape and crystallization condition. The shape
of Lip 42 crystal appeared to be partly attributed to the
presence of N-terminal tag. |
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