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10_1007_s00449-017-1757-3 | chunk_1 | Bioprocess Biosyst Eng CrossMark DOI 10.1007/s00449-017-1757-3 RESEARCH PAPER Interactions of phosphate solubilising microorganisms with natural rare-earth phosphate minerals: a study utilizing Western Australian monazite Melissa K. Corbett1 · Jacques J. Eksteen? · Xi-Zhi Niu3 · Jean-Philippe Croue3 . Elizabeth L. J. W... |
10_1007_s00449-017-1757-3 | chunk_2 | The application of microbial leaching is a proven fungi using the Lysing matrix A tube (MP Biomedi- technology for the recovery of copper, nickel, zinc, and cals） with 500 μL of lysis buffer [25] in a ribolyser cobalt from low-grade ores [22]. It is a well-established (MP Biomedicals) for 30 s at a speed of 4.0. RNase ... |
10_1007_s00449-017-1757-3 | chunk_3 | Nota- concentrations of P were corrected for decreases in fuid bly, all three fungal species demonstrated complete clear- volume due to evaporation and the sampling process. ing of the media with large pellets formed. Of the bacte- Separation and identification of organic acids produced rial species tested, K. pneumoni... |
10_1007_s00449-017-1757-3 | chunk_4 | After 24 h with CSM, all bacterial species demon- cies when grown on CSM was greater than when grown on strated a decrease in pH to below 4.6, finishing between 2.6 MWM; however, there was a greater variation in the range and 4.12 by 192 h (192 h data depicted in Fig. 7). Fungal of organic acids produced by the bacteri... |
10_1007_s00449-017-1757-3 | chunk_5 | In addition, during the solvent extrac- with Nd levels approximately 87-97% of those recorded tion process, thorium is co-precipitated along with the for La. Incubation with B. megaterium and Ps. aeruginosa REEs [38] resulting in contamination of the leach liquor. leached Nd at levels 20% greater than La, even though t... |
10_1007_s00449-017-1757-3 | chunk_6 | It is likely that unidentified acids in unaffected by increases in soluble Mn levels. this study were important in solubilising the MWM result- Numerous strains of B. megaterium are known to solu- ing in the higher REE levels detected and lower pH in the bilise phosphate [66] and are actively applied in agriculture lea... |
10_1007_s00449-017-1757-3 | chunk_7 | Lett Appl Microbiol 30:53-56 processing, recycling and associated environmental issues. 26. White TJ, Bruns T, Lee S, Taylor J (1990) Amplification and National Risk Management Research Laboratory direct sequencing of fungal ribosomal RNA genes for phyloge- 3. Chu S (2011) Critical materials strategy. In: Energy UDo (e... |
10_1007_s00449-017-1757-3 | chunk_8 | Clark DS, Ito K, Horitsu H (1966) Effect of manganese and other schaften 91:552-555 heavy metals on submerged citric acid fermentation of molasses. 70. Schneider KD, Van Straaten P, De Orduna RM, Glasauer, S, Tre- Biotechnol Bioeng 8:465-471 vors J, Fallow D, Smith PS (2010) Comparing phosphorus mobi- 53. Acharya C, Ka... |
10_1006_icar_2002_6837 | chunk_1 | Icarus 158, 98-105 (2002) doi:10.1006/icar.2002.6837 Hidden Mass in the Asteroid Belt G. A. Krasinsky, E. V. Pitjeva, M. V. Vasilyev, and E. I. Yagudina Institute ofApplied Astronomy,Russian Academy of Sciences,Kutuzov Quay 10,St.Petersburg 191187,Russia E-mail: kra @ quasar.ipa.nw.ru Received November 13, 2001; revise... |
10_1006_icar_2002_6837 | chunk_2 | Fortunately it ap- of the main belt (which have become a routine procedure-at pears possible to derive rather accurate masses of these asteroids present 30 asteroids of the main belt have been measured in this in the process of fitting the planetary ephemerides to the ranging way; see Magri et al. 1999). Comparison wit... |
10_1006_icar_2002_6837 | chunk_3 | In fact the SBN database keeps two NEAR sets of radi: a set derived from IRAS observations (they are re- 0.00003362 0.00051938 ferred hereafter as the “"radiometric radi' or “system 1 of radi') IRAS 0.000058 0.000698 and a larger set from the IMPS Ground Data File ("system 2"). HIDDEN MASS IN THE ASTEROID BELT 101 So, ... |
10_1006_icar_2002_6837 | chunk_4 | In Fig. 3 both the curve of this distribution and the 20 km. The typical mass of asteroids from the last subset is about experimental data (for about 2000 minor planets) are depicted. 0.0001 × 10-10 M。 with the total mass ≈1 x 10-10 M。 whose HIDDEN MASS IN THE ASTEROID BELT 103 value is not negligible if one processes ... |
10_1006_icar_2002_6837 | chunk_5 | Mainbelt asteroids: Results of Arecibo and Goldstone radar observa- tions of 37 objects during 1980-1995. Icarus 140, 379-407. Michalak, G. 2000. Determination of asteroid masses. I. (1) Ceres, (2) Pallas, Then for the perturbing force of the ring we obtain and (4) Vesta. Astron. Astrophys. 360, 363-374. Michalak, G. 2... |
10_1016_j_mineng_2005_10_024 | chunk_1 | Available online at www.sciencedirect.com SCIENCI )DIRECT MINERALS ENGINEERING ELSEVIER Minerals Engineering 19 (2006) 960-967 This article is also available online at: www.elsevier.com/locate/mineng Formation of jarosite during Fe2+ oxidation by Acidithiobacillus ferrooxidans J. Daoud, D. Karamanev * Department of Che... |
10_1016_j_mineng_2005_10_024 | chunk_2 | Furthermore, jarosite formation in Smith et al. (1988) 2.0-2.3 immobilization matrices limits the amount of biomass Drobner et al. (1990) 2.0 retention since ferric iron deposits occupy most of the Optimum temperature (°C) available space (Jensen and Webb, 1995). Ahonen and Tuovinen (1989) 28 Okereke and Stevens (1991)... |
10_1016_j_mineng_2005_10_024 | chunk_3 | Several experimental runs were performed testing the effect of the various pHs, range from 1.0 to 3.0 in 0.2 inter- vals, taking samples initially, at 22 h and at 46 h. The tem- oxidation progression with time for the various pHs are perature was kept constant at 22 °C. The observations of given in Table 2. 0.14 0.14 0... |
10_1016_S0168-1656_02_00222-5 | chunk_1 | ELSEVIER Journal of Biotechnology 99 (2002) 319-330 www.elsevier.com/locate/jbiotec MELISSA: a loop of interconnected bioreactors to develop life support in Space F. Godia a*, J. Albiola, J.L. Montesinosa, J. Péreza, N. Creus ?, F. Cabello a, X. Menguala, A. Montras a, Ch. Lasseur b aDepartament d'Enginyeria Quimica,Es... |
10_1016_S0168-1656_02_00222-5 | chunk_2 | The continuous long term ments, a medium is defined as a combination of operation of the nitrification reactor is done at both, on the basis that all the compounds are different oxygen levels, since this is a critical present at the required level. parameter for this reaction. The bioreactor for compartment IVa has bee... |
10_1016_S0168-1656_02_00222-5 | chunk_3 | Fi- the steady-state operation of the bioreactor has nally, the correlation of Fr with the voltage been characterized, basically in terms of ammoni- applied to the lamps allowed to obtain the final um degradation efficiency at varying loads (ob- calibration curve. tained by changing either t the ammonium concentration ... |
10_1016_S0168-1656_02_00222-5 | chunk_4 | A final photo- model is fully described in the literature (Cornet et bioreactor of 77 1 volume was designed according al., 1992a,b, 1998), and it is used in the control of to various criteria: maintaining cylindrical geome- the continuous operation of the bioreactor. Indeed try, maximizing the ratio between illuminated... |
10_1016_S0168-1656_02_00222-5 | chunk_5 | From this point, approach to reach the final construction of the the oxygen level is consecutively lowered to 20 loop. The experiments reported in this contribu- (period V), 15 (period VI), 10 (period VII), 5% tion demonstrate that the long term continuous (period VIII), and finally raised back to 40% operation of thes... |
ijms-26-01905-v3 | references | Citation: Fukumoto, Y.; Li, E.;
Tanaka, Y.; Suzuki, N.; Ogra, Y.
Evaluation of Metal Accumulation in
Escherichia coli Expressing SPL2 by
Single-Cell Inductively Coupled
Plasma Mass Spectrometry. Int. J. Mol.
Sci.2025 ,26, 1905. https://doi.org/
10.3390/ijms26051905
Copyright: © 2025 by the authors.
Licensee MDPI, Basel... |
ijms-26-01905-v3 | experiments | Evaluation of Metal Accumulation in Escherichia coli Expressing
SPL2 by Single-Cell Inductively Coupled Plasma
Mass Spectrometry
Yasunori Fukumoto1,*
, Enhui Li1, Yu-ki Tanaka1
, Noriyuki Suzuki1,2and Yasumitsu Ogra1
1Graduate School of Pharmaceutical Sciences, Chiba University, Chiba 260-8675, Japan;
yu-ki.tanaka@chib... |
ijms-26-01905-v3 | abstract | Abstract: Rare earth elements, comprising 17 elements including 15 lanthanides, are es-
sential components in numerous high-tech applications. While physicochemical methods
are commonly employed to remove toxic heavy metals (e.g., cadmium and mercury) from
industrial wastewater, biological approaches offer increasingly... |
ijms-26-01905-v3 | discussion | resulted in high expression levels and solubility. Single-cell ICP-MS analysis revealed that
these recombinant bacteria accumulated lanthanum, cobalt, nickel, and cadmium, effec-
tively sequestering lanthanum and cadmium from the culture media. Furthermore, SPL2 |
ijms-26-01905-v3 | results | expression conferred enhanced bacterial tolerance to cadmium exposure. These findings
establish SPL2 as a promising candidate for developing recombinant bacterial systems for
heavy metal bioremediation and rare earth element biomining.
Keywords: scICP-MS; single-cell inductively coupled plasma mass spectrometry; SPL2;
... |
ijms-26-01905-v3 | introduction | 1. Introduction
Rare earth elements (REEs), a group of 17 chemical elements including 15 lanthanides,
yttrium, and scandium [ 1], are indispensable in numerous high-tech applications owing to
their unique physicochemical properties [ 2–4]. However, conventional REE mining gener-
ates substantial amounts of REE-rich liq... |
ijms-26-01905-v3 | methodology | offers a more sustainable approach. Advances in molecular and genetic engineering have
further enhanced the efficiency of REE recovery in biomining processes. A key molecular
engineering strategy involves genetically modifying microorganisms to express specific
proteins, such as lanthanide-binding tags, which significa... |
ijms-26-01905-v3 | methodology | used for heavy metal removal, these methods are often hampered by significant limitations,
including high reagent consumption and the generation of hazardous sludges [18].
Bioremediation, utilizing microorganisms to adsorb and transport metal ions into their |
ijms-26-01905-v3 | methodology | cells via active transport [ 19], offers an eco-friendly alternative to conventional methods
for addressing heavy metal pollution. Naturally occurring bacteria, fungi, seaweeds, and
algae have demonstrated significant potential for heavy metal adsorption and accumula-
tion [ 20–23], offering advantages such as superior... |
ijms-26-01905-v3 | discussion | dividual cells. scICP-MS offers several advantages over bulk analysis using conventional
ICP-MS. Namely, acid decomposition is not required, reducing the potential risks of contam-
Int. J. Mol. Sci. 2025 ,26, 1905 3 of 13
ination and sample loss. In addition, inaccuracies in cell counting may affect quantification |
ijms-26-01905-v3 | discussion | accuracy in a bulk analysis. scICP-MS overcomes these limitations by enabling the direct
analysis of individual cells. Moreover, it provides a more comprehensive characterization
of samples by showing not only the average elemental content but also the variability
and distribution patterns within the population [ 34–38... |
ijms-26-01905-v3 | methodology | required for in vitro studies. This approach facilitated the characterization of the metal-
binding properties of recombinant proteins [ 39], demonstrating the power of scICP-MS for
cellular metalloproteomics.
Of the three lanthanide-binding proteins investigated in this study, SPL2 demonstrated
superior characteristic... |
ijms-26-01905-v3 | results | 2. Results
2.1. SPL2 and Lanpepsy Show High Expression and Solubility in Bacteria
To examine the expression of potential lanthanoid-binding proteins, we expressed
SPL2 (residues V291–S383) fused with flag-mCherry in E. coli . Robust expression was
observed for both SPL2-flag-mCherry and lanpepsy-flag (Figure 1A). Howev... |
ijms-26-01905-v3 | discussion | binding ability [ 39], further analysis focused on soluble proteins SPL2-flag-mCherry and
lanpepsy-flag. To confirm the solubility of the recombinant proteins, we analyzed the
soluble fraction of bacterial lysates. Sufficient amounts of SPL2-flag-mCherry and lanpepsy-
flag were detected (Figure 1B), demonstrating their... |
ijms-26-01905-v3 | discussion | dividual cells. scICP-MS o ffers several advantages over bulk analysis using conventional
ICP-MS. Namely, acid decomposition is not required, reducing the potential risks of con-
tamination and sample loss. In addition, inaccuracies in cell counting may a ffect quanti fi-
cation accuracy in a bulk an alysis. scICP-MS ove... |
ijms-26-01905-v3 | discussion | direct analysis of individual cells. Moreover , it provides a more comprehensive character-
ization of samples by showing not only the average elemental content but also the varia-
bility and distribution pa tterns within the population [34–38]. In a recent study, we em-
ployed scICP-MS to directly detect histidin e-ta... |
ijms-26-01905-v3 | methodology | steps typically required for in vitro studies. This approach facilitated the characterization
of the metal-binding properties of recombinan t proteins [39], demonstrating the power of
scICP-MS for cellular metalloproteomics.
Of the three lanthanide-binding proteins investigated in this study, SPL2 demon-
strated sup... |
ijms-26-01905-v3 | results | 2. Results
2.1. SPL2 and Lanpepsy Show High Expression and Solubility in Bacteria
To examine the expression of potential la nthanoid-binding proteins, we expressed
SPL2 (residues V291 ‒S383) fused with flag-mCherry in E. coli . Robust expression was ob-
served for both SPL2- flag-mCherry and lanpepsy- flag (Figure 1A).... |
ijms-26-01905-v3 | discussion | metal-binding ability [39], further analysis focused on soluble proteins SPL2- flag-
mCherry and lanpepsy- flag. To con firm the solubility of the recombinant proteins, we
analyzed the soluble fraction of bacterial lysates. Su fficient amounts of SPL2- flag-mCherry
and lanpepsy- flag were detected (Figure 1B), demonstrating... |
ijms-26-01905-v3 | results | statistically significant increase in La binding compared with control groups. These results
demonstrate that the SPL2 fragment facilitates enhanced La binding.
While SPL2 expression tended to increase Tb binding, this difference was not statisti-
cally significant (Figure 3). The flag-mCherry-expressing group also sho... |
ijms-26-01905-v3 | results | statistically signi ficant increase in La binding compared with control groups. These results
demonstrate that the SPL2 fragment facilitates enhanced La binding.
While SPL2 expression tended to increase Tb binding, this di fference was not statis-
tically signi ficant (Figure 3). The flag-mCherry-expressing group also sh... |
ijms-26-01905-v3 | experiments | tribution of La signal intensities. Representati ve results from three independent experiments are
shown, with Gaussian distribution fitting curves overlaid. ( F) The amount of La per bacterial cell
was calculated from scICP-MS signal intensities, and the average content per cell was determined. |
ijms-26-01905-v3 | experiments | The graph shows data from thr ee independent experiments. *, p < 0.05. ***, p < 0.001.
Figure 2. Lanthanum (La) binding in SPL2-expressing bacteria. ( A–E) Recombinant bacteria were
cultured in liquid medium, and recombinant protein expression was induced. SPL2-flag-mCherry
and control proteins were expressed and then... |
ijms-26-01905-v3 | experiments | of La signal intensities. Representative results from three independent experiments are shown, with
Gaussian distribution fitting curves overlaid. ( F) The amount of La per bacterial cell was calculated
from scICP-MS signal intensities, and the average content per cell was determined. The graph shows |
ijms-26-01905-v3 | discussion | Figure 3. Terbium (Tb) binding by recombinant bacteria. ( A‒E) Tb binding analysis by scICP-MS.
The experimental procedure was identical to that de scribed in Figure 2, except that the recombinant
bacteria were exposed to 250 µM Tb ion. ( F) The graph shows the averag e amount of bound Tb per |
ijms-26-01905-v3 | experiments | cell. Data represent means from th ree independent experiments. *, p < 0.05.
2.3. SPL2 Enhances Transition Metal Binding in Recombinant Bacteria
The ability of SPL2- flag-mCherry-expressing bacteria to bind transition metals was
analyzed. Previously, we demonstrated that re combinant bacteria expressing 6His-tagged
... |
ijms-26-01905-v3 | experiments | cell. Data represent means from three independent experiments. *, p< 0.05.
Int. J. Mol. Sci. 2025 ,26, 1905 5 of 13
2.3. SPL2 Enhances Transition Metal Binding in Recombinant Bacteria
The ability of SPL2-flag-mCherry-expressing bacteria to bind transition metals was
analyzed. Previously, we demonstrated that recombinan... |
ijms-26-01905-v3 | discussion | Figure 3. Terbium (Tb) binding by recombinant bacteria. ( A‒E) Tb binding analysis by scICP-MS.
The experimental procedure was identical to that de scribed in Figure 2, except that the recombinant
bacteria were exposed to 250 µM Tb ion. ( F) The graph shows the averag e amount of bound Tb per |
ijms-26-01905-v3 | experiments | cell. Data represent means from th ree independent experiments. *, p < 0.05.
2.3. SPL2 Enhances Transition Metal Binding in Recombinant Bacteria
The ability of SPL2- flag-mCherry-expressing bacteria to bind transition metals was
analyzed. Previously, we demonstrated that re combinant bacteria expressing 6His-tagged
... |
ijms-26-01905-v3 | experiments | of bound Ni per cell. Data represent means from four independent experiments. *, p< 0.05.
Int. J. Mol. Sci. 2025 , 26, x FOR PEER REVIEW 6 of 14
Figure 5. Cobalt (Co) binding by recombinant bacteria expressing mCherry-6His and SPL2- flag- |
ijms-26-01905-v3 | experiments | bound Co per cell. Data represent means from three independent experiments. *, p < 0.05.
The Cd binding of SPL2-expressing bacter ia was also examined. SPL2 expression
caused a rightward shift in the histogram (Figure 6). The SPL2- flag-mCherry-expressing
group showed a signi ficant increase in Cd binding compared wit... |
ijms-26-01905-v3 | results | These results indicate that the SPL2 fragment within the SPL2- flag-mCherry protein pro-
moted Cd binding of the recombinant bacteria.
Figure 6. Cadmium (Cd) binding by recombin ant bacteria expressing SPL2- flag-mCherry. ( A‒E) |
ijms-26-01905-v3 | experiments | The experimental procedure was identical to that de scribed in Figure 2, with the exception that the
recombinant bacteria were exposed to 250 µM Cd ion. ( F) The graph shows the average amount of
bound Cd per cell. Data represent means from three independent experiments. *, p < 0.05. **, p < 0.01.
2.4. La and Cd Seq... |
ijms-26-01905-v3 | experiments | bound Co per cell. Data represent means from three independent experiments. *, p< 0.05.
The Cd binding of SPL2-expressing bacteria was also examined. SPL2 expression
caused a rightward shift in the histogram (Figure 6). The SPL2-flag-mCherry-expressing
Int. J. Mol. Sci. 2025 ,26, 1905 6 of 13
group showed a significant... |
ijms-26-01905-v3 | results | These results indicate that the SPL2 fragment within the SPL2-flag-mCherry protein pro-
moted Cd binding of the recombinant bacteria.
Int. J. Mol. Sci. 2025 , 26, x FOR PEER REVIEW 6 of 14
Figure 5. Cobalt (Co) binding by recombinant bacteria expressing mCherry-6His and SPL2- flag- |
ijms-26-01905-v3 | experiments | bound Co per cell. Data represent means from three independent experiments. *, p < 0.05.
The Cd binding of SPL2-expressing bacter ia was also examined. SPL2 expression
caused a rightward shift in the histogram (Figure 6). The SPL2- flag-mCherry-expressing
group showed a signi ficant increase in Cd binding compared wit... |
ijms-26-01905-v3 | results | These results indicate that the SPL2 fragment within the SPL2- flag-mCherry protein pro-
moted Cd binding of the recombinant bacteria.
Figure 6. Cadmium (Cd) binding by recombin ant bacteria expressing SPL2- flag-mCherry. ( A‒E) |
ijms-26-01905-v3 | experiments | The experimental procedure was identical to that de scribed in Figure 2, with the exception that the
recombinant bacteria were exposed to 250 µM Cd ion. ( F) The graph shows the average amount of
bound Cd per cell. Data represent means from three independent experiments. *, p < 0.05. **, p < 0.01.
2.4. La and Cd Seq... |
ijms-26-01905-v3 | experiments | bound Cd per cell. Data represent means from three independent experiments. *, p< 0.05. **, p< 0.01.
2.4. La and Cd Sequestration in SPL2-Expressing Bacteria
We evaluated the ability of recombinant bacteria to remove Cd and La from the
culture medium. When cultured in La-containing medium, La concentrations in the cul-... |
ijms-26-01905-v3 | results | Consistent with the La removal results, Cd concentration in the supernatant showed
a signi ficant reduction when the SPL2-expressin g bacteria were cultured in media con-
taining 250 µM and 25 µM Cd (Figure 7D ‒F). The SPL2-expressing group also exhibited a
tendency for enhanced Cd removal at 50 µM Cd. These findings s... |
ijms-26-01905-v3 | experiments | Graphs show means from thr ee independent experiments. p-values were calculated using Student’s
t-test or Welch’s t-test, comparing each bacterial group with the no-bacteria control (culture medium
without bacteria), except for pa nel F. 6His, mCherry-6His; FLAG, flag-mCherry; SPL2, SPL2- flag-
mCherry. *, p < 0.05. **... |
ijms-26-01905-v3 | results | Consistent with the La removal results, Cd concentration in the supernatant showed
a signi ficant reduction when the SPL2-expressin g bacteria were cultured in media con-
taining 250 µM and 25 µM Cd (Figure 7D ‒F). The SPL2-expressing group also exhibited a
tendency for enhanced Cd removal at 50 µM Cd. These findings s... |
ijms-26-01905-v3 | experiments | Graphs show means from thr ee independent experiments. p-values were calculated using Student’s
t-test or Welch’s t-test, comparing each bacterial group with the no-bacteria control (culture medium
without bacteria), except for pa nel F. 6His, mCherry-6His; FLAG, flag-mCherry; SPL2, SPL2- flag-
mCherry. *, p < 0.05. **... |
ijms-26-01905-v3 | results | Co, La, Tb, or Ni (Figure 8B,C). These results strongly suggest that SPL2 expression
Figure 7. Removal of La and Cd ions from culture media by SPL2-expressing bacteria. ( A–C) La
removal. ( D–F) Cd removal. Recombinant bacteria were cultured in liquid medium, and recombinant
protein expression was induced. Bacteria we... |
ijms-26-01905-v3 | experiments | MS. Graphs show means from three independent experiments. p-values were calculated using
Student’s t-test or Welch’s t-test, comparing each bacterial group with the no-bacteria control (culture
medium without bacteria), except for panel F. 6His, mCherry-6His; FLAG, flag-mCherry; SPL2,
SPL2-flag-mCherry. *, p< 0.05. **,... |
ijms-26-01905-v3 | results | Consistent with the La removal results, Cd concentration in the supernatant showed a
significant reduction when the SPL2-expressing bacteria were cultured in media containing
250µM and 25 µM Cd (Figure 7D–F). The SPL2-expressing group also exhibited a tendency |
ijms-26-01905-v3 | results | for enhanced Cd removal at 50 µM Cd. These findings suggest that SPL2-expressing
bacteria effectively sequester Cd from the culture medium.
2.5. SPL2 Expression Increases Cd Tolerance of Recombinant Bacteria
We examined the impact of SPL2 expression on bacterial tolerance to lanthanides and
transition metals. SPL2-expr... |
ijms-26-01905-v3 | results | Co, La, Tb, or Ni (Figure 8B,C). These results strongly suggest that SPL2 expression specifically
confers tolerance to Cd, but not to the other tested metals, in the recombinant bacteria.
Int. J. Mol. Sci. 2025 , 26, x FOR PEER REVIEW 8 of 14
specifically confers tolerance to Cd, but not to the other tested metals, i... |
ijms-26-01905-v3 | experiments | tive control. Representative results from more than two independent experiments are shown. Con-
trol images for Cd and Co are identical. In panel C, pairs of the control and SPL2 images were taken
from the same image. |
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