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Characterization of the algC gene expression pattern in the multidrug resistant Acinetobacter baumannii AIIMS 7 and correlation with biofilm development on abiotic surface.

Sahu PK, Iyer PS, Barage SH, Sonawane KD, Chopade BA - ScientificWorldJournal (2014)

Bottom Line: Comparison revealed differential algC expression pattern with maximum 81.59-fold increase in biofilm cells versus 3.24-fold in planktonic cells (P < 0.05).Expression levels strongly correlated with specific biofilm stages (scale of 3 to 96 h), coinciding maximum at initial surface attachment stage (9 h) and biofilm maturation stage (48 h).Moreover, molecular dynamics analysis on the three-dimensional structure of PMM/PGM (simulated up to 10 ns) revealed enzyme structure as stable and similar to that in P. aeruginosa (synthesis of alginate and lipopolysaccharide core) and involved in constitution of biofilm EPS (extracellular polymeric substances).

View Article: PubMed Central - PubMed

Affiliation: Institute of Bioinformatics and Biotechnology, University of Pune, Pune 411 007, India ; Ispat General Hospital, SAIL, Rourkela 769 005, India.

ABSTRACT
Relative quantification of algC gene expression was evaluated in the multidrug resistant strain Acinetobacter baumannii AIIMS 7 biofilm (3 to 96 h, on polystyrene surface) compared to the planktonic counterparts. Comparison revealed differential algC expression pattern with maximum 81.59-fold increase in biofilm cells versus 3.24-fold in planktonic cells (P < 0.05). Expression levels strongly correlated with specific biofilm stages (scale of 3 to 96 h), coinciding maximum at initial surface attachment stage (9 h) and biofilm maturation stage (48 h). Cloning, heterologous expression, and bioinformatics analyses indicated algC gene product as the bifunctional enzyme phosphomannomutase/phosphoglucomutase (PMM/PGM) of ∼ 53 kDa size, which augmented biofilms significantly in algC clones compared to controls (lacking algC gene), further localized by scanning electron microscopy. Moreover, molecular dynamics analysis on the three-dimensional structure of PMM/PGM (simulated up to 10 ns) revealed enzyme structure as stable and similar to that in P. aeruginosa (synthesis of alginate and lipopolysaccharide core) and involved in constitution of biofilm EPS (extracellular polymeric substances). Our observation on differential expression pattern of algC having strong correlation with important biofilm stages, scanning electron-microscopic evidence of biofilm augmentation taken together with predictive enzyme functions via molecular dynamic (MD) simulation, proposes a new basis of A. baumannii AIIMS 7 biofilm development on inanimate surfaces.

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SDS-PAGE analysis of AlgC (PMM/PGM) protein. Lane 1: whole cell extract of control E. coli DH5α (lacking algC gene); lane 2: whole cell extract of E. coli DH5α with algC gene showing the AlgC protein band ~53 kDa; lane M: standard protein molecular weight marker (kDa).
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fig5: SDS-PAGE analysis of AlgC (PMM/PGM) protein. Lane 1: whole cell extract of control E. coli DH5α (lacking algC gene); lane 2: whole cell extract of E. coli DH5α with algC gene showing the AlgC protein band ~53 kDa; lane M: standard protein molecular weight marker (kDa).

Mentions: To demonstrate the synthesis of PMM/PGM protein, whole cell extracts of cloned E. coli DH5α with resultant recombinant plasmid pGEalgCA7 cells were used in SDS-PAGE analysis, which showed a clearly overexpressed protein of ~53 kDa suggesting the PMM/PGM protein in question (Figure 5). Control E. coli DH5α (with empty vector pGEM-3Zf+ lacking an algC gene) did not show any PMM/PGM protein band.


Characterization of the algC gene expression pattern in the multidrug resistant Acinetobacter baumannii AIIMS 7 and correlation with biofilm development on abiotic surface.

Sahu PK, Iyer PS, Barage SH, Sonawane KD, Chopade BA - ScientificWorldJournal (2014)

SDS-PAGE analysis of AlgC (PMM/PGM) protein. Lane 1: whole cell extract of control E. coli DH5α (lacking algC gene); lane 2: whole cell extract of E. coli DH5α with algC gene showing the AlgC protein band ~53 kDa; lane M: standard protein molecular weight marker (kDa).
© Copyright Policy - open-access
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC4269089&req=5

fig5: SDS-PAGE analysis of AlgC (PMM/PGM) protein. Lane 1: whole cell extract of control E. coli DH5α (lacking algC gene); lane 2: whole cell extract of E. coli DH5α with algC gene showing the AlgC protein band ~53 kDa; lane M: standard protein molecular weight marker (kDa).
Mentions: To demonstrate the synthesis of PMM/PGM protein, whole cell extracts of cloned E. coli DH5α with resultant recombinant plasmid pGEalgCA7 cells were used in SDS-PAGE analysis, which showed a clearly overexpressed protein of ~53 kDa suggesting the PMM/PGM protein in question (Figure 5). Control E. coli DH5α (with empty vector pGEM-3Zf+ lacking an algC gene) did not show any PMM/PGM protein band.

Bottom Line: Comparison revealed differential algC expression pattern with maximum 81.59-fold increase in biofilm cells versus 3.24-fold in planktonic cells (P < 0.05).Expression levels strongly correlated with specific biofilm stages (scale of 3 to 96 h), coinciding maximum at initial surface attachment stage (9 h) and biofilm maturation stage (48 h).Moreover, molecular dynamics analysis on the three-dimensional structure of PMM/PGM (simulated up to 10 ns) revealed enzyme structure as stable and similar to that in P. aeruginosa (synthesis of alginate and lipopolysaccharide core) and involved in constitution of biofilm EPS (extracellular polymeric substances).

View Article: PubMed Central - PubMed

Affiliation: Institute of Bioinformatics and Biotechnology, University of Pune, Pune 411 007, India ; Ispat General Hospital, SAIL, Rourkela 769 005, India.

ABSTRACT
Relative quantification of algC gene expression was evaluated in the multidrug resistant strain Acinetobacter baumannii AIIMS 7 biofilm (3 to 96 h, on polystyrene surface) compared to the planktonic counterparts. Comparison revealed differential algC expression pattern with maximum 81.59-fold increase in biofilm cells versus 3.24-fold in planktonic cells (P < 0.05). Expression levels strongly correlated with specific biofilm stages (scale of 3 to 96 h), coinciding maximum at initial surface attachment stage (9 h) and biofilm maturation stage (48 h). Cloning, heterologous expression, and bioinformatics analyses indicated algC gene product as the bifunctional enzyme phosphomannomutase/phosphoglucomutase (PMM/PGM) of ∼ 53 kDa size, which augmented biofilms significantly in algC clones compared to controls (lacking algC gene), further localized by scanning electron microscopy. Moreover, molecular dynamics analysis on the three-dimensional structure of PMM/PGM (simulated up to 10 ns) revealed enzyme structure as stable and similar to that in P. aeruginosa (synthesis of alginate and lipopolysaccharide core) and involved in constitution of biofilm EPS (extracellular polymeric substances). Our observation on differential expression pattern of algC having strong correlation with important biofilm stages, scanning electron-microscopic evidence of biofilm augmentation taken together with predictive enzyme functions via molecular dynamic (MD) simulation, proposes a new basis of A. baumannii AIIMS 7 biofilm development on inanimate surfaces.

Show MeSH
Related in: MedlinePlus