Biochemical localization of a protein involved in synthesis of Gluconacetobacter hansenii cellulose

Iyer, Prashanti R.; Catchmark, Jeffrey M.; Brown, Nicole R.

doi:10.1007/s10570-011-9504-4

Cited by 21 publications

(16 citation statements)

References 35 publications

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“…One of the proteins coded by the operon, AcsD, has been reported to have a cylindershaped octamer structure localized in the periplasmic space, and it has been suggested that this structure may play a role in cellulose crystallization by functioning Patterns of cell movement include advancement in one direction rotation and then advancement in the other direction (b); oscillation side-to-side (c); spinning about axes (d); single directional non-linear advancement (e); and nonordered reversals (f) as a part of the cellulose synthesizing enzyme complex (Saxena et al 1994;Hu et al 2010;Iyer et al 2011). The purpose of this study was to characterize the disruption mutant to aid in determining the function of AcsD.…”

Section: Discussionmentioning

confidence: 99%

“…In G. xylinus, normal arrangement of the linear terminal complexes appears as a single row along the longitudinal axis of the cell (Kimura et al 2001). The AcsD octamer was determined to be localized in the same linear arrangement within the cell's periplasm (Hu et al 2010;Iyer et al 2011;Sunagawa et al 2013). The linear TC orientation on the mutant cells did not have this same linear arrangement; instead, the linear TCs appeared abnormal as they were often observed to be positioned on the transverse, diagonal, or longitudinal axis (Figs.…”

Section: Acsd Characterizationmentioning

confidence: 99%

“…The structure and arrangement of the AcsD protein itself indicates a cylinder with the presence of a functional complex unit in the form of an octamer (Hu et al 2010). The octamer is localized in the periplasm with an orientation parallel to the long axis of the cell where it interacts with CcpAx (Hu et al 2010;Iyer et al 2011;Sunagawa et al 2013). Additionally, it has an interior space that may act as a channel through which a nascent glucan chain may pass through, thus supporting the concept that it may be a part of the pore complex; however, its true function is still unknown (Hu et al 2010;Iyer et al 2011).…”

Section: Introductionmentioning

confidence: 99%

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Characterization of an acsD disruption mutant provides additional evidence for the hierarchical cell-directed self-assembly of cellulose in Gluconacetobacter xylinus

2014

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The acsD gene is involved in cellulose biosynthesis among the Acetobacter species. In the current study, we created an acsD disruption mutant in the acsABCD cellulose synthase operon of Gluconacetobacter xylinus and characterized the resulting cellulose to aid in providing insight into the function of the acsD gene. Both the wild type G. xylinus AY201 (derivative of Gluconacetobacter hansenii ATCC 23769) and the acsD disruption mutant produced crystalline cellulose I microfibrils. The cellulose produced by both appeared to be synthesized from an aggregate of pores known as a linear terminal complex; however the total cellulose synthesized was 10 % that of the wild type G. xylinus AY201. TEM observations of the acsD disruption mutant confirmed that microfibrils and bundles of microfibrils were similar in size to the G. xylinus AY201 wild type; however, the final ribbon dimensions were narrower (53.4 ± 13.1 nm wt, vs. 28.2 ± 8.2 nm). Additional TEM observations of the mutant cells incubated at 4°C revealed an abnormal linear terminal complex orientation whereby the resulting band material could be observed in a transverse orientation as well as longitudinally to the long axis of the cell. Taken together, these data strongly suggest that acsD aids in the proper orientation of the linear terminal complexes along the longitudinal axis of the cell indicating the AcsD protein is involved in the final level of the hierarchical assembly of cellulose resulting in highly efficient cellulose synthesis.

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Section: Discussionmentioning

confidence: 99%

Section: Acsd Characterizationmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

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Characterization of an acsD disruption mutant provides additional evidence for the hierarchical cell-directed self-assembly of cellulose in Gluconacetobacter xylinus

2014

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“…The other protein we examined for impact on cellulose synthesis activity was AcsD, which is a periplasmic localized protein [32].…”

Section: The Catalytic Core Of the Cscmentioning

confidence: 99%

AcsA–AcsB: The core of the cellulose synthase complex from Gluconacetobacter hansenii ATCC23769

McManus

Deng

Nagachar

et al. 2016

Enzyme and Microbial Technology

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The gram-negative bacterium, Gluconacetobacter hansenii, produces cellulose of exceptionally high crystallinity in comparison to the cellulose of higher plants. This bacterial cellulose is synthesized and extruded into the extracellular medium by the cellulose synthase complex (CSC). The catalytic component of this complex is encoded by the gene AcsAB. However, several other genes are known to encode proteins critical to cellulose synthesis and are likely components of the bacterial CSC. We have purified an active heterodimer AcsA-AcsB from G. hansenii ATCC23769 to homogeneity by two different methods. With the purified protein, we have determined how it is post-translationally processed, forming the active heterodimer AcsA-AcsB. Additionally, we have performed steady-state kinetic studies on the AcsA-AcsB complex. Finally through mutagenesis studies, we have explored the roles of the postulated CSC proteins AcsC, AcsD, and CcpAx.

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“…AcsD is the only Acs protein of Gluconacetobacter whose crystal structure has been determined (11); it is an octameric protein assembly localized to the periplasmic space (11,12). It may function as a cellulose chaperone, directing the newly synthesized cellulose chains to AcsC in the outer membrane to prevent accumulation of rogue strands in the periplasm.…”

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confidence: 99%

Identification and Characterization of Non-Cellulose-Producing Mutants of Gluconacetobacter hansenii Generated by Tn 5 Transposon Mutagenesis

et al. 2013

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The acs operon of Gluconacetobacter is thought to encode AcsA, AcsB, AcsC, and AcsD proteins that constitute the cellulose synthase complex, required for the synthesis and secretion of crystalline cellulose microfibrils. A few other genes have been shown to be involved in this process, but their precise role is unclear. We report here the use of Tn5 transposon insertion mutagenesis to identify and characterize six non-cellulose-producing (Cel ؊ ) mutants of Gluconacetobacter hansenii ATCC 23769. The genes disrupted were acsA, acsC, ccp Ax (encoding cellulose-complementing protein [the subscript "Ax" indicates genes from organisms formerly classified as Acetobacter xylinum]), dgc1 (encoding guanylate dicyclase), and crp-fnr (encoding a cyclic AMP receptor protein/fumarate nitrate reductase transcriptional regulator). Protein blot analysis revealed that (i) AcsB and AcsC were absent in the acsA mutant, (ii) the levels of AcsB and AcsC were significantly reduced in the ccp Ax mutant, and (iii) the level of AcsD was not affected in any of the Cel ؊ mutants. Promoter analysis showed that the acs operon does not include acsD, unlike the organization of the acs operon of several strains of closely related Gluconacetobacter xylinus. Complementation experiments confirmed that the gene disrupted in each Cel ؊ mutant was responsible for the phenotype. Quantitative real-time PCR and protein blotting results suggest that the transcription of bgl Ax (encoding ␤-glucosidase and located immediately downstream from acsD) was strongly dependent on Crp/Fnr. A bgl Ax knockout mutant, generated via homologous recombination, produced only ϳ16% of the wild-type cellulose level. Since the crp-fnr mutant did not produce any cellulose, Crp/Fnr may regulate the expression of other gene(s) involved in cellulose biosynthesis.

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Biochemical localization of a protein involved in synthesis of Gluconacetobacter hansenii cellulose

Cited by 21 publications

References 35 publications

Characterization of an acsD disruption mutant provides additional evidence for the hierarchical cell-directed self-assembly of cellulose in Gluconacetobacter xylinus

Characterization of an acsD disruption mutant provides additional evidence for the hierarchical cell-directed self-assembly of cellulose in Gluconacetobacter xylinus

AcsA–AcsB: The core of the cellulose synthase complex from Gluconacetobacter hansenii ATCC23769

Identification and Characterization of Non-Cellulose-Producing Mutants of Gluconacetobacter hansenii Generated by Tn 5 Transposon Mutagenesis

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