Extracellular Synthesis, Specific Recognition, and Intracellular Degradation of Cyclomaltodextrins by the Hyperthermophilic Archaeon
            <i>Thermococcus</i>
            sp. Strain B1001

Hashimoto, Yoshiteru; Yamamoto, Tomoko; Fujiwara, Shinsuke; Takagi, Masahiro; Imanaka, Tadayuki

doi:10.1128/jb.183.17.5050-5057.2001

Cited by 67 publications

(57 citation statements)

References 51 publications

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“…Specifically, PF0478 shows 64 and 39% sequence identity with the extracellular cyclomaltodextrin glucanotransferases (CGTases) of Thermococcus kodakaraensis KOD1 (35) and Thermococcus sp. strain B1001 (19), respectively. Surprisingly, PF0478 is not predicted to contain a signal sequence by the SignalP, TargetP, and SOSUI programs.…”

Section: Resultsmentioning

confidence: 99%

Transcriptional and Biochemical Analysis of Starch Metabolism in the Hyperthermophilic Archaeon Pyrococcus furiosus

et al. 2006

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Pyrococcus furiosus utilizes starch and its degradation products, such as maltose, as primary carbon sources, but the pathways by which these ␣-glucans are processed have yet to be defined. For example, its genome contains genes proposed to encode five amylolytic enzymes (including a cyclodextrin glucanotransferase [CGTase] and amylopullulanase), as well as two transporters for maltose and maltodextrins (Mal-I and Mal-II), and a range of intracellular enzymes have been purified that reportedly metabolize maltodextrins and maltose. However, precisely which of these enzymes are involved in starch processing is not clear. In this study, starch metabolism in P. furiosus was examined by biochemical analyses in conjunction with global transcriptional response data for cells grown on a variety of glucans. In addition, DNA sequencing led to the correction of two key errors in the genome sequence, and these change the predicted properties of amylopullulanase (now designated PF1935*) and CGTase (PF0478*). Based on all of these data, a pathway is proposed that is specific for starch utilization that involves one transporter (Mal-II [PF1933 to PF1939]) and only three enzymes, amylopullulanase (PF1935*), 4-␣-glucanotransferase (PF0272), and maltodextrin phosphorylase (PF1535). Their expression is upregulated on starch, and together they generate glucose and glucose-1-phosphate, which then feed into the novel glycolytic pathway of this organism. In addition, the results indicate that several hypothetical proteins encoded by three gene clusters are also involved in the transport and processing of ␣-glucan substrates by P. furiosus.Pyrococcus furiosus is an obligately anaerobic, heterotrophic, hyperthermophilic archaeon that was isolated from a shallow hydrothermal vent near Vulcano Island, Italy (12). The organism grows optimally near 100°C and is able to utilize a range of sugars as a primary carbon source (4,11,28,29). These include cellobiose (28, 43), laminarin (43), chitin (16), maltose (29), barley glucan (3), and starch (29). However, the means by which these compounds are metabolized by P. furiosus are not well understood. In attempting to define the pathway by which starch, and one of its degradation products, maltose, serve as carbon sources, there are two complicating factors. First, the genome sequence of P. furiosus (36) contains an array of genes that encode putative glycosyl hydrolases and glycosyl transferases. With respect to starch, they include five enzymes that potentially hydrolyze ␣-glucans. Two of them (PF0477 and PF1935) have putative signal peptides and are presumably extracellular, while the other three (PF0272, PF0478, and PF1939) lack a signal peptide and are presumably intracellular. They are annotated as amylases (PF0272 and PF0477), amylopullulanases (PF1934 and PF1935), and cyclodextrin glucanotransferases (PF0478). The question is, are all of these enzymes involved in degrading starch? The pathway for utilizing maltose and maltodextrins, primary products of starch breakdown, is well defined in ...

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Section: Resultsmentioning

confidence: 99%

Transcriptional and Biochemical Analysis of Starch Metabolism in the Hyperthermophilic Archaeon Pyrococcus furiosus

et al. 2006

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“…Second, the open reading frames for CDase, cyclomaltodextrin glucanotransferase, and the CD uptake system are commonly found as a gene cluster on the chromosomal DNA in several organisms, including Klebsiella oxytoca (21), Thermococcus kodakaraensis KOD1 (22), and Thermococcus sp. strain B1001 (23). Cyclomaltodextrin glucanotransferase is an extracellular enzyme, whereas CDase (MAase and NPase) is an intracellular enzyme.…”

Section: Discussionmentioning

confidence: 99%

Cyclomaltodextrinase, Neopullulanase, and Maltogenic Amylase Are Nearly Indistinguishable from Each Other

Lee¹,

Kim²,

Cho³

et al. 2002

Journal of Biological Chemistry

147

124

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Over 20 enzymes denoted as cyclomaltodextrinase, maltogenic amylase, or neopullulanase that share 40 -86% sequence identity with each other are found in public data bases. These enzymes are distinguished from typical ␣-amylases by containing a novel N-terminal domain and exhibiting preferential substrate specificities for cyclomaltodextrins (CDs) over starch. In this research field, a great deal of confusion exists regarding the features distinguishing the three groups of enzymes from one another. Although a different enzyme code has been assigned to each of the three different enzyme names, even a single differentiating enzymatic property has not been documented in the literature. On the other hand, an outstanding question related to this issue concerns the structural basis for the preference of these enzymes for CDs. To clarify the confusion and to address this question, we have determined the structures of two enzymes, one from alkalophilic Bacillus sp. I-5 and named cyclomaltodextrinase and the other from a Thermus species and named maltogenic amylase. The structure of the Bacillus enzyme reveals a dodecameric assembly composed of six copies of the dimer, which is the structural and functional unit of the Thermus enzyme and an enzyme named neopullulanase. The structure of the Thermus enzyme in complex with ␤-CD led to the conclusion that Trp 47 , a well conserved N-terminal domain residue, contributes greatly to the preference for ␤-CD. The common dimer formation through the novel N-terminal domain, which contributes to the preference for CDs by lining the active-site cavity, convincingly indicates that the three groups of enzymes are not different enough to preserve the different names and enzyme codes.

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“…B1001 (30), maltogenic amylase from Thermus sp. IM6501 (14), neopullulanase from B. stearothermophilus (31), and T. vulgaris ␣-amylase II (13).…”

Section: Methodsmentioning

confidence: 99%

Association of Novel Domain in Active Site of Archaic Hyperthermophilic Maltogenic Amylase from Staphylothermus marinus

Jung

Park

et al. 2012

Journal of Biological Chemistry

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Background: Maltogenic amylases that are known to date form dimers to perform hydrolysis. Results: The structure of maltogenic amylase from Staphylothermus showed a novel domain at the N terminus associated with the active site. Conclusion:Staphylothermus amylase has all of its substrate-binding structural components in a single monomer. Significance: This is the first report of the newly observed domain arrangement adopted by hyperthermophilic archaic maltogenic amylase.

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Extracellular Synthesis, Specific Recognition, and Intracellular Degradation of Cyclomaltodextrins by the Hyperthermophilic Archaeon Thermococcus sp. Strain B1001

Cited by 67 publications

References 51 publications

Transcriptional and Biochemical Analysis of Starch Metabolism in the Hyperthermophilic Archaeon Pyrococcus furiosus

Transcriptional and Biochemical Analysis of Starch Metabolism in the Hyperthermophilic Archaeon Pyrococcus furiosus

Cyclomaltodextrinase, Neopullulanase, and Maltogenic Amylase Are Nearly Indistinguishable from Each Other

Association of Novel Domain in Active Site of Archaic Hyperthermophilic Maltogenic Amylase from Staphylothermus marinus

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