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The Prokaryotic Microalga Limnothrix redekei KNUA012 to Improve Aldehyde Decarbonylase Expression for Use as a Biological Resource

Young-Saeng Kim
Young-Saeng Kim
, 
Haeri Baek
Haeri Baek
, 
Hyun-Sik Yun
Hyun-Sik Yun
, 
Jae-Hak Lee
Jae-Hak Lee
, 
Kyoung-In Lee
Kyoung-In Lee
, 
Han-Soon Kim
Han-Soon Kim
 und 
Ho-Sung Yoon
Ho-Sung Yoon
   | 20. Sept. 2023
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Article Category: Original Paper
Online veröffentlicht: 20. Sept. 2023
Seitenbereich: 307 - 317
Eingereicht: 15. März 2023
Akzeptiert: 04. Juli 2023
DOI: https://doi.org/10.33073/pjm-2023-031
Schlüsselwörter
© 2023 Young-Saeng Kim et al., published by Sciendo
This work is licensed under the Creative Commons Attribution-NonCommercial-NoDerivatives 4.0 International License.

Fig. 1.

Phylogenetic relationships of Limnothrix redekei KNUA012 and related organisms.The tree is constructed to scale with branch lengths in the same units as those of the evolutionary distances used to infer the phylogenetic tree. The evolutionary distances were computed using the maximum composite likelihood method (Tamura et al. 2004), and evolutionary analyses were conducted using MEGA5 (Tamura et al. 2011).
Phylogenetic relationships of Limnothrix redekei KNUA012 and related organisms.The tree is constructed to scale with branch lengths in the same units as those of the evolutionary distances used to infer the phylogenetic tree. The evolutionary distances were computed using the maximum composite likelihood method (Tamura et al. 2004), and evolutionary analyses were conducted using MEGA5 (Tamura et al. 2011).

Fig. 2.

a) Light microscopy images of Limnothrix redekei KNUA012. The cell diameter ranged from 10 μm depending on the growth stage. Live cells were visualized at magnifications of 400 × (left panel) and 1,000 × (right panel) under a microscope equipped with differential interference contrast optics.b) Growth curves of Limnothrix redekei KNUA012 at 5–35°C (at intervals of 5°C).
a) Light microscopy images of Limnothrix redekei KNUA012. The cell diameter ranged from 10 μm depending on the growth stage. Live cells were visualized at magnifications of 400 × (left panel) and 1,000 × (right panel) under a microscope equipped with differential interference contrast optics.b) Growth curves of Limnothrix redekei KNUA012 at 5–35°C (at intervals of 5°C).

Fig. 3.

SDS polyacrylamide gel images of the aldehyde decarbonylase (AD) protein purified from Escherichia coli cells transformed with the expression vector pET-28a carrying AD.a) M – marker, lane 1 – protein purified from E. coli cells transformed with pET-28a (pET28 empty vector), lane 2 – protein purified from E. coli cells transformed with pET28-AD (pET-28a harboring the AD gene); b) M – marker, lane 3 – purified AD obtained from E. coli (pET28-AD).
SDS polyacrylamide gel images of the aldehyde decarbonylase (AD) protein purified from Escherichia coli cells transformed with the expression vector pET-28a carrying AD.a) M – marker, lane 1 – protein purified from E. coli cells transformed with pET-28a (pET28 empty vector), lane 2 – protein purified from E. coli cells transformed with pET28-AD (pET-28a harboring the AD gene); b) M – marker, lane 3 – purified AD obtained from E. coli (pET28-AD).

Fig. 4.

Analysis of the GC peaks.a) GC/MS total ion chromatogram of standard fatty acid methyl esters. 1 – Tridecanoic acid methyl ester, 2 – pentadecanoic acid methyl ester, 3 – heptadecanoic acid methyl ester, 4 – nonadecanoic acid methyl ester, and 5 – henelcosanoic acid methyl ester.b) GC peak results for the fatty acids extracted from L. redekei KNUA012. 1 – Pentadecane, 2 – dodecanoic acid methyl ester, 3 – 8-heptadecene, 4 – heptadecane, 5 – methyl Z-11-tetradecenoate, 6 – tetradecanoic acid methyl ester, 7 – 9-hexadecenoic acid methyl ester, 8 – palmitoleic acid methyl ester, 9 – hexadecanoic acid methyl ester, 10 – 9-octadecenoic acid methyl ester, and 11 – octadecanoic acid methyl ester. Peak no. annotated in Table SI
Analysis of the GC peaks.a) GC/MS total ion chromatogram of standard fatty acid methyl esters. 1 – Tridecanoic acid methyl ester, 2 – pentadecanoic acid methyl ester, 3 – heptadecanoic acid methyl ester, 4 – nonadecanoic acid methyl ester, and 5 – henelcosanoic acid methyl ester.b) GC peak results for the fatty acids extracted from L. redekei KNUA012. 1 – Pentadecane, 2 – dodecanoic acid methyl ester, 3 – 8-heptadecene, 4 – heptadecane, 5 – methyl Z-11-tetradecenoate, 6 – tetradecanoic acid methyl ester, 7 – 9-hexadecenoic acid methyl ester, 8 – palmitoleic acid methyl ester, 9 – hexadecanoic acid methyl ester, 10 – 9-octadecenoic acid methyl ester, and 11 – octadecanoic acid methyl ester. Peak no. annotated in Table SI

GC/MS results showing the alkanes and major fatty acids present in Limnothrix redekei KNUA012, Escherichia coli cells transformed with pET28-AD, and the empty vector pET28.

Peak no. Component name pET28 empty vector (% w/w) KNUA012 (% w/w) pET28-AD (% w/w)
1 Pentadecane 0.87 ± 0.52 2.40 ± 0.85 1.78 ± 0.38
2 Dodecanoic acid methyl ester 0.18 ± 0.05 0.54 ± 0.21 0.25 ± 0.08
3 8-Heptadecene 0.54 ± 0.07 0.62 ± 0.23 1.09 ± 0.31
4 Heptadecane 1.07 ± 0.17 1.15 ± 0.42 1.86 ± 0.52
5 Methyl Z-11-tetradecenoate 21.7 ± 1.52 22.1 ± 1.03 23.6 ± 1.59
6 Tetradecanoic acid methyl ester 11.2 ± 1.31 11.2 ± 1.07 15.7 ± 1.41
7 9-Hexadecenoic acid methyl ester 2.62 ± 0.84 21.3 ± 1.51 8.63 ± 1.52
8 Palmitoleic acid methyl ester 2.35 ± 0.53 5.37 ± 1.03 3.82 ± 0.82
9 Hexadecanoic acid methyl ester 2.06 ± 0.51 21.7 ± 1.85 7.82 ± 1.31
10 9-Octadecenoic acid methyl ester 3.64 ± 0.85 9.31 ± 1.37 6.83 ± 1.46
11 Octadecanoic acid methyl ester 0.84 ± 0.06 1.04 ± 0.52 1.83 ± 0.82
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