Zentrum für Ultrastrukturforschung und Ludwig Bolzmann-Institut für Molekulare Nanotechnologie, Universität für Bodenkultur, A-1180 Vienna, Austria1
Author for correspondence: Eva M. Egelseer. Tel: +43 1 47 654 2233. Fax: +43 1 478 91 12. e-mail: egelseer{at}edv1.boku.ac.at
![]() |
ABSTRACT |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
Keywords: insertion-sequence element, S-layer protein, Bacillus stearothermophilus
Abbreviations: DIG-dUTP, digoxigenin-11-dUTP; IS, insertion sequence; S-layer, surface layer
The GenBank accession number for the sequence reported in this paper is AF162268.
![]() |
INTRODUCTION |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
Bacillus stearothermophilus is a Gram-positive, strictly aerobic species of endospore-forming bacteria that can produce large amounts of exoenzymes such as proteases and amylases (Archibald, 1989 ; Priest, 1981
). In previous reports, the putative role of the S-layers from B. stearothermophilus DSM 2358 and ATCC 12980 (DSM 22) as an adhesion site for a high-molecular-mass amylase was confirmed (Egelseer et al., 1995
, 1996
). To gather information as to whether S-layer lattices of Gram-positive bacteria delineate a kind of periplasmic space and may control the speed of exoenzyme release, comparative studies were carried out with the S-layer-carrying (S+) strain of B. stearothermophilus ATCC 12980 and its S-layer-deficient (S-) derivative (Egelseer et al., 1996
). The S- strain was isolated from SVIII agar slants of the S+ strain which were stored for 2 years at 4 °C (Eder, 1983
). In freeze-etched preparations, a highly ordered S-layer lattice with oblique symmetry could only be observed on whole cells from the S+ strain, whereas the S- strain revealed an amorphous cell surface, typical of strains lacking an S-layer (Egelseer et al., 1996
).
Recently, the nucleotide sequence encoding the S-layer protein SbsC from the S+ strain of B. stearothermophilus ATCC 12980 was determined by PCR techniques (Jarosch et al., 2000 ). The entire sbsC sequence shows an ORF of 3297 bp predicted to encode a protein of 1099 aa (AF055578) with a theoretical molecular mass of 115409 Da and a pI of 5·73. Primer-extension analysis indicated the existence of two promoter regions which are most probably active in different growth stages (Jarosch et al., 2000
).
The loss in S-layer protein synthesis has been described for several organisms during prolonged cultivation under optimal laboratory conditions, indicating that these crystalline arrays provide a selective advantage in competitive natural habitats (Messner & Sleytr, 1992 ). On the other hand, environmental stress factors such as temperature upshift also caused a loss in S-layer-protein synthesis (Sleytr et al., 1982
). The first detailed studies to understand the molecular mechanism leading to the loss in S-layer protein synthesis were carried out with Aeromonas salmonicida, a fish-pathogenic organism. It was demonstrated that the insertion of insertion sequence (IS) elements at different positions of the vapA gene encoding the S-layer (A-layer) protein and in its upstream region had occurred (Gustafson et al., 1994
).
In general, IS elements are small translocating segments of DNA that are located on the host chromosome and on plasmids with the potential to move within and among bacterial genomes. While these elements have a variety of architectures, there are two essential features common to all transposable elements. Firstly, they are delineated by end sequences that are required in cis for the transposition reaction. Secondly, they encode the functions that facilitate and control their movement. Gene inactivation or macrogenomic rearrangements such as adjacent deletions, inversions and cointegrate formations are often discovered as a result of ISs (Gasson & Fitzgerald, 1993 ). Today over 500 bacterial ISs isolated from 73 genera representing 159 species of bacteria and archaea have been characterized at the nucleotide-sequence level (Mahillon & Chandler, 1998
). It is now evident that transposition does not occur randomly, since in each case studied, some target preference for integration has been observed (Craig, 1997
). The integration of an IS element can either repress or activate genes located downstream.
In the present study, we report the sequence determination and the characterization of a novel type of bacterial IS element, designated ISBst12. This IS element was found to be responsible for the loss of S-layer protein synthesis in the S- strain of B. stearothermophilus ATCC 12980.
![]() |
Methods |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
DNA manipulations and oligonucleotides used for PCR.
Chromosomal DNA of the S+ and S- strains of B. stearothermophilus ATCC 12980 was prepared by using Genomic Tips 100 (Qiagen) according to the manufacturers instructions. Digestion of DNA with restriction endonucleases and separation of DNA fragments by agarose gel electrophoresis were performed as described by Sambrook et al. (1989) . DNA fragments were recovered from agarose gels by using the Qiaex II Gel Extraction Kit (Qiagen). Restriction endonucleases were purchased from Roche Molecular Biochemicals. All oligonucleotides were synthesized on an Applied Biosystems DNA synthesizer.
Detection and PCR amplification of ISBst12.
The presence of the sbsC gene sequence was investigated in the S- strain by using the primer combinations sbsC3/12, sbsC3/14, sbsC14/22, sbsC21/22 and sbsC12/19 (Table 1). PCR reactions were performed in a 50 µl reaction volume containing 240 µM each dNTP, 240 nM primer, 1·25 mM MgCl2, 1 U Taq DNA polymerase (Roche Molecular Biochemicals) and 100 ng chromosomal DNA from either the S+ or S- strain of B. stearothermophilus ATCC 12980 as template in 1xTaq reaction buffer (Roche Molecular Biochemicals) according to the manufacturers instructions. Thirty cycles of amplification were performed in a thermocycler (Hybaid Touch Down Control). Each cycle consisted of a 30 s denaturation step at 95 °C, a 45 s annealing step with annealing temperatures depending on the calculated Tm of the oligonucleotides used and extension times of 60 s per 1000 bp at 72 °C. PCR fragments generated in at least four separate reaction vials were pooled and the mixture was subjected to DNA sequencing. For investigation of the occurrence of ISBst12 in B. stearothermophilus PV72/p6, PV72/p2 and PV72/T5, PCR reactions were carried out as described above, except that 1 µl of the suspensions of exponentially growing cells were used as the template. The primer combination ISBst12-1/2 was used to detect ISBst12 in these organisms (Table 1
). As a control, the S+ and S- strains of B. stearothermophilus ATCC 12980 were included into the study.
|
Isolation of RNA.
Total RNA was isolated from cultures of exponentially growing cells of the S+ and S- strains of B. stearothermophilus ATCC 12980 which had reached an OD600 of 0·78. For enzymic lysis, the cell pellet obtained by centrifugation of 4·5 ml bacterial suspension at 5000 g for 5 min at 4 °C was resuspended in 0·5 ml 10 mM Tris/HCl buffer, pH 8·0, containing 1 mM EDTA and 5 mg lysozyme (Sigma) ml1. Samples were incubated for 15 min at room temperature. Isolation of total RNA was performed by using the RNeasy Midi Kit (Qiagen) according to the manufacturers instructions. The RNA concentration was determined spectrophotometrically at 260 nm and samples were stored at -20 °C.
DNA and RNA hybridization.
Northern and Southern blotting was performed as described by Sambrook et al. (1989) . Southern blots were carried out with chromosomal DNA from the S+ and S- strains of B. stearothermophilus ATCC 12980 digested with either HindIII or SalI. For hybridization of Southern blots, a DNA probe generated by using the primer combination ISBst12-1/2 was labelled with digoxigenin-11-dUTP (DIG-dUTP) by incorporation during PCR. Hybridization was performed according to the manufacturers recommendations. For Northern blotting, total RNA from the S+ and S- strains (5 µg per well) was fractionated by electrophoresis through a 1% agarose/0·22 M formaldehyde gel. Hybridization of Northern blots was carried out either with the ISBst12-specific DNA probe or with a 3·3 kb DNA fragment of the sbsC gene generated by the primer pair sbsC3/12 and randomly labelled with DIG-dUTP. Hybrids were detected using the DIG Luminescent Detection Kit (Roche Molecular Biochemicals). The size of the transcripts was estimated from their mobilities relative to those of the RNAs in the DIG-dUTP labelled RNA Molecular Mass Marker II (size range 1·56·9 kb; Roche Molecular Biochemicals).
![]() |
RESULTS |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
Localization and sequence characterization of ISBst12
By using the sbsC-specific primer combinations sbsC3/12, sbsC3/14 and sbsC14/22, the PCR-generated fragments of the sbsC gene from the S- strain increased in size by approximately 1600 bp compared to that from the S+ strain (Fig. 1, lanes 2, 4 and 6). However, by using the sbsC-derived primer combinations sbsC21/22 and sbsC12/19, comprising the upstream and the downstream region of the sbsC gene, respectively, the PCR-generated fragments from the S+ and S- strains were the same size (Fig. 1
, lanes 710). These preliminary results obtained from PCR indicated that the coding region of the sbsC gene in the S- strain carried an insertion of approximately 1600 bp. The nucleotide sequence determination of the insertion was carried out on an 1800 bp PCR fragment generated from genomic DNA of the S- strain by using the primer combination sbsC3/14 (Fig. 1
, lane 4). The insertion site of the foreign DNA was found to be located within the coding region of the sbsC gene, 199 bp downstream from the ATG start codon. Due to the insertion of the foreign sequence, a stop codon was introduced into the ORF of the sbsC gene. Nucleotide-sequence analysis revealed that the insertion was 1612 bp long, bounded by 16 bp imperfect inverted repeats and flanked by a directly repeated 8 bp target sequence originating from the sbsC gene. This DNA sequence displayed the structural features of bacterial IS elements and thus was designated ISBst12 (accession no. AF162268). The G+C content of ISBst12 was determined to be 47%. The IS element contained one ORF of 1446 bp, predicted to encode a protein of 482 aa. The ORF started with ATG, preceded by a typical prokaryotic ribosome-binding site (AAGGAGG).
|
|
|
|
|
|
![]() |
DISCUSSION |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
As revealed by Southern blotting, ISBst12 was present in the genome of the S- strain as well as in the S+ strain in multiple copies, which was also described for Lactococcus lactis harbouring at least 12 copies of IS905 (Dodd et al., 1994 ). The hybridization patterns, which were slightly different between the S+ and S- strains, indicated the insertion of at least two more copies of ISBst12 elsewhere in the genome of the S- strain. Furthermore, an ISBst12-specific transcript could exclusively be detected in the S- strain. Taken together, these results could indicate that an active transposase is only present in the S- strain, leading to an increase in ISBst12 copy number compared to the S+ strain in which transcription of ISBst12 seems to be inhibited or considerably reduced. Furthermore, by using PCR, ISBst12 was also detected in B. stearothermophilus PV72/p6, its oxygen-induced strain variant PV72/p2 and the S-layer-deficient strain PV72/T5 (Sára et al., 1996
). The latter also carries the IS element IS4712, which was found to be responsible for the loss of S-layer gene expression (Scholz, 1998
).
The putative transposase encoded by ISBst12 has a calculated pI of 9·13, which is consistent with a probable DNA-binding property required for transposase activity. The N-terminal region of the transposase displays two overlapping leucine-zipper motifs (Landschulz et al., 1988 ). These heptad repeats of leucines were originally described as a protein-dimerization motif for the formation of coiled-coil intertwining of
-helices for several eukaryotic transcriptional regulators and also for some prokaryotic DNA-binding proteins like MetR (Maxon et al., 1990
) and
54 (Sasse-Dwight & Gralla, 1990
). Interestingly, leucine-zipper motifs were also detected in many members of the IS3 family, for example in IS2 (Lei & Hu, 1997
), IS911 (Haren et al., 1997
) and IS1221 (Zheng & McIntosh, 1995
). This is consistent with the observation that some, if not all transposases, including prokaryotic and eukaryotic elements such as retroviruses, have the capacity to generate multimeric forms essential for their activity (Polard & Chandler, 1995
). In the putative transposase encoded by ISBst12, a highly basic region was identified adjacent to the leucine-zipper motif that is characteristic of eukaryotic DNA-binding proteins but was absent in the prokaryotic DNA-binding proteins MetR and
54 as well as in IS1221 (Zheng & McIntosh, 1995
). From the increasing number of different transposable elements isolated and characterized at the nucleotide-sequence level, a general pattern for the functional organization is emerging, namely that the sequence-specific DNA-binding activities are located at the N-terminal region, while the catalytic domain is mostly located at the C-terminal end (Mahillon & Chandler, 1998
). One functional interpretation of this arrangement for prokaryotic elements is that it may permit the interaction of a nascent polypeptide chain with its target sequences on the IS, thus coupling expression and activity. This notion is reinforced by the observation that the presence of the C-terminal region of several transposases [IS50 (Weinreich et al., 1993
), IS10 (Jain & Kleckner, 1993
)] appears to mask the DNA-binding domain and decreases the binding activity, thereby favouring the activity of the protein in cis. This preferential activity in cis reduces the probability that transposase expression from a given element would activate transposition of related copies elsewhere in the genome. This could also be true of the putative transposase encoded by ISBst12 carrying a highly acidic region towards the C-terminal end which after complete folding of the polypeptide chain probably masks the basic DNA-binding region located at the N-terminal part.
Comparative studies between bacterial transposases and retroviral integrases revealed similarities in a region that is thought to form part of the active site, namely the DDE motif (Polard & Chandler, 1995 ). This highly conserved acidic amino acid triad was found to be intimately involved in catalysis by coordinating divalent metal cations (in particular Mg2+) implicated in assisting the various nucleophilic attacking groups during the course of the reaction. The majority of the IS elements found in the genus Bacillus were assigned to the IS4 family (Mahillon & Chandler, 1998
). This family is quite heterogeneous but it is characterized by a conserved DDE signature (Rezsöhazy et al., 1993
). However, the putative transposase encoded by ISBst12 does not exhibit a typical DDE triad. Although members of the DDE family represent the majority of known IS elements, and mutagenic studies clearly underlined the importance of these residues, a significant fraction of IS elements does not exhibit a real or potential DDE triad. Interestingly, a highly conserved His-Arg-Tyr triad was identified in the putative transposase which resembles the signature of the catalytic site of integrases of the bacteriophage
Int family and was also detected in the C-terminal part of the IS1 transposase (Abremski & Hoess, 1992
; Argos et al., 1986
; Serre et al., 1995
). For the IS1 transposase it was demonstrated that each of the three amino acid residues of the conserved triad is important for transposase activity (Serre et al., 1995
).
While no sequence identities to IS elements were identified at the nucleotide-sequence level, the protein encoded by ISBst12 revealed identity to several putative transposases encoded by bacterial IS elements. The transposase encoded by ISBst12 showed the highest identity value (35%) to a recently identified putative transposase located on a 46 kb plasmid of D. radiodurans (White et al., 1999 ). This extremely radiation-resistant bacterium contains numerous insertion sequences (52 copies) and small non-coding repeats (247 copies) whose evolutionary significance and role in genome function remain unclear (White et al., 1999
; Makarova et al., 1999
). Furthermore, a gas-vesicle protein (ORF H0698) that was detected in both large inverted repeats identified on the Halobacterium sp. plasmid pNRC 100 (Ng et al., 1991
) revealed identity to the putative ISBst12 transposase. These two large inverted repeats were found to mediate inversion of the intervening single-copy region (Ng et al., 1991
). The transposase encoded by ISBst12 showed identity to a putative transposase encoded by IS22-1 of V. cholerae as well as to two members of the IS66 family, namely to a protein encoded by IS66 of A. tumefaciens and to a protein encoded by ISRm14 of S. meliloti. Like ISBst12, members of the IS66 family are flanked by 8 bp direct target repeats and by terminal inverted repeats of 1527 bp, which are very similar among members of this family. Both 16 bp imperfect inverted repeats of ISBst12 start with 5'-GTAA-3', a sequence which seems to be conserved among the inverted repeats of several members from the IS66 family (Mahillon & Chandler, 1998
). The IS66 family was found to be restricted to agrobacteria and rhizobia (Mahillon & Chandler, 1998
). With the development of studies on the mechanism of bacterial pathogenesis, an association between IS elements and many pathogenic and virulence functions became evident. Such associations have been observed in animal pathogens like Vibrio, in plant pathogens like Agrobacterium or in symbionts like Rhizobium.
To conclude, ISBst12 represents a novel type of IS element, whose putative transposase exhibits a well defined leucine-zipper motif as well as a His-Arg-Tyr triad and reveals sequence identity to transposases from different IS families detected in distantly related genera of bacteria (Deinococcus, Vibrio, Agrobacterium, Sinorhizobium) and even to a plasmid-encoded halobacterial protein. Since it could be speculated that this novel type of IS element may represent an ancestral IS, the distribution and abundance among bacterial, archaeal and even eukaryotic genera remains to be investigated.
![]() |
ACKNOWLEDGEMENTS |
---|
![]() |
REFERENCES |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
Altschul, S. F., Madden, T. L., Schäffer, A. A., Zhang, J., Zhang, Z., Miller, W. & Lipman, D. J. (1997). Gapped BLAST and PSI-BLAST: a new generation of protein database search programs. Nucleic Acids Res 25, 3389-3402.
Archibald, R. (1989). The Bacillus cell envelope. In Bacillus: Biotechnology Handbook 2 , pp. 217-254. Edited by C. R. Harwood. New York: Plenum.
Argos, P., Landy, A., Abremski, K. & 9 other authors (1986). The integrase family of site-specific recombinases: regional similarity and global diversity. EMBO J 5, 433440.[Abstract]
Bartelmus, W. & Perschak, F. (1957). Schnellmethode zur Keimzahlbestimmung in der Zuckerindustrie. Z Zuckerind 7, 276-281.
Craig, N. (1997). Target site selection in transposition. Annu Rev Biochem 66, 437-474.[Medline]
Dodd, H. M., Horn, N. & Gasson, M. J. (1994). Characterization of IS905, a new multicopy insertion sequence identified in Lactococci. J Bacteriol 176, 3393-3396.[Abstract]
Eder, J. (1983). Versuche zur Aufklärung der Funktion parakristalliner Proteinmembranen bei Bacillus stearothermophilus. PhD thesis, University of Agricultural Sciences, Vienna.
Egelseer, E. M., Schocher, I., Sára, M. & Sleytr, U. B. (1995). The S-layer from Bacillus stearothermophilus DSM 2358 functions as an adhesion site for a high-molecular-weight amylase. J Bacteriol 177, 14441451.[Abstract]
Egelseer, E. M., Schocher, I., Sleytr, U. B. & Sára, M. (1996). Evidence that an N-terminal S-layer protein fragment triggers the release of a cell-associated high-molecular-weight amylase from Bacillus stearothermophilus ATTC 12980. J Bacteriol 178, 5602-5609.
Gasson, M. J. & Fitzgerald, G. F. (1993). Gene-tranfer systems and transposition. In Genetics and Biotechnology of Lactic Acid Bacteria , pp. 1-51. Edited by M. J. Gasson & W. M. De Vos. Glasgow: Blackie Academic and Professional.
Gustafson, C. E., Chu, S. & Trust, T. (1994). Mutagenesis of the paracrystalline surface protein array of Aeromonas salmonicida by endogenous insertion elements. J Mol Biol 237, 452-463.[Medline]
Haren, L., Bétermier, M., Polard, P. & Chandler, M. (1997). IS911-mediated intramolecular transposition is naturally temperature sensitive. Mol Microbiol 25, 531-540.[Medline]
Jain, C. & Kleckner, N. (1993). Preferential cis action of IS10 transposase depends upon its mode of synthesis. Mol Microbiol 9, 249-260.[Medline]
Jarosch, M., Egelseer, E. M., Mattanovich, D., Sleytr, U. B. & Sára, M. (2000). S-layer gene sbsC of Bacillus stearothermophilus ATCC 12980: molecular characterization and heterologous expression in Escherichia coli. Microbiology 146, 273-281.
Kleckner, N. (1990). Regulation of transposition in bacteria. Annu Rev Cell Biol 6, 297-327.
Laemmli, U. K. (1970). Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature 227, 680-685.[Medline]
Landschulz, W. H., Johnson, P. F. & McKnight, S. L. (1988). The leucine zipper: a hypothetical structure common to a new class of DNA-binding proteins. Science 240, 1759-1764.[Medline]
Lei, G.-S. & Hu, S.-T. (1997). Functional domains of InsA protein of IS2. J Bacteriol 179, 6238-6243.[Abstract]
Machida, Y., Sakurai, M., Kiyokawa, S., Ubasawa, A., Suzuki, Y. & Ikeda, J. E. (1984). Nucleotide sequence of the insertion sequence found in the T-DNA region of mutant Ti plasmid pTiA66 and distribution of its homologues in octopine Ti plasmid. Proc Natl Acad Sci USA 81, 7495-7499.[Abstract]
Mahillon, J. & Chandler, M. (1998). Insertion sequences. Microbiol Mol Biol Rev 62, 725-774.
Makarova, K. S., Wolf, Y. I., White, O., Minton, K. & Daly, M. J. (1999). Short repeats and IS elements in the extremely radiation-resistant bacterium Deinococcus radiodurans and comparison to other species. Res Microbiol 150, 711-724.[Medline]
Maxon, M. E., Wigboldus, J., Brot, N. & Weissbach, H. (1990). Structure-function studies on Escherichia coli MetR protein, a putative prokaryotic leucine zipper protein. Proc Natl Acad Sci USA 87, 7076-7079.[Abstract]
Messner, P. & Sleytr, U. B. (1992). Crystalline bacterial cell surface layers. Adv Microb Physiol 33, 213-275.[Medline]
Ng, W. L., Kothakota, S. & DasSarma, S. (1991). Structure of the gas vesicle plasmid in Halobacterium halobium: inversion isomers, inverted repeats, and insertion sequences. J Bacteriol 173, 1958-1964.[Medline]
Polard, P. & Chandler, M. (1995). Bacterial transposases and retroviral integrases. Mol Microbiol 15, 13-23.[Medline]
Priest, F. G. (1981). Products and applications. In Bacillus: Biotechnology Handbooks 2 , pp. 293-320. Edited by C. R. Harwood. New York: Plenum.
Rezsöhazy, R., Hallet, B., Delcour, J. & Mahillon, J. (1993). The IS4 family of insertion sequences: evidence for a conserved transposase motif. Mol Microbiol 9, 1283-1295.[Medline]
Rost, B. & Sander, C. (1993). Prediction of protein structure at better than 70% accuracy. J Mol Biol 232, 584-599.[Medline]
Sambrook, J., Fritsch, E. F. & Maniatis, T. (1989). Molecular Cloning: a Laboratory Manual, 2nd edn. Cold Spring Harbor, NY: Cold Spring Harbor Laboratory.
Sára, M. & Sleytr, U. B. (2000). S-layer proteins. J Bacteriol 182, 859-868.
Sára, M., Kuen, B., Mayer, H. F., Mandl, F., Schuster, K. C. & Sleytr, U. B. (1996). Dynamics in oxygen-induced changes in S-layer protein synthesis from Bacillus stearothermophilus PV72 and its S-layer deficient variant T5 in continuous culture and studies on the cell-wall composition. J Bacteriol 178, 2108-2117.[Abstract]
Sasse-Dwight, S. & Gralla, J. D. (1990). Role of eukaryotic-type functional domains found in the prokaryotic enhancer receptor factor 54. Cell 62, 945-954.[Medline]
Schneiker, S., Kosier, B., Puehler, A. & Selbitschka, W. (1999). The Sinorhizobium meliloti insertion sequence (IS) element ISRm14 is related to a previously unrecognized IS element located adjacent to the Escherichia coli locus of enterocyte effacement (LEE) pathogenicity island. Curr Microbiol 39, 274-281.[Medline]
Scholz, H. (1998). Genetische Analyse der S-layer Protein Variation in Bacillus stearothermophilus PV72. PhD thesis, University of Vienna.
Serre, M.-C., Turlan, C., Bortolin, M.-L. & Chandler, M. (1995). Mutagenesis of the IS1 transposase: importance of the His-Arg-Tyr triad for activity. J Bacteriol 177, 5070-5077.[Abstract]
Sleytr, U. B. & Sára, M. (1997). Bacterial and archaeal S-layer proteins: structure-function relationships and their biotechnological applications. Trends Biotechnol 15, 20-26.[Medline]
Sleytr, U. B., Messner, P., Pum, D. & Eder, J. (1982). Struktur und morphogenese periodischer proteinmembranen bei bakterien. Mikroskopie 39, 215-232.[Medline]
Sleytr, U. B., Messner, P., Pum, D. & Sára, M. (1993). Crystalline bacterial cell surface layers. Mol Microbiol 10, 911-916.[Medline]
Sleytr, U. B., Messner, P., Pum, D. & Sára, M. (1999). Crystalline bacterial cell surface layers (S-layers): from cell structure to biomimetics and nanotechnology. Angew Chem Int Ed Engl 38, 1034-1054.
Weinreich, M. D., Mahnke-Braam, L. & Reznikoff, W. S. (1993). A functional analysis of the Tn5 transposase: identification of domains required for DNA binding and multimerization. J Mol Biol 241, 166-177.
White, O., Eisen, J. A., Heidelberg, J. F. & 29 other authors (1999). Genome sequence of the radioresistant bacterium Deinococcus radiodurans R1. Science 286, 15711577.
Yamasaki, S., Shimizu, T., Hoshino, R., Ho, S.-T., Shimada, T., Nair, G. B. & Takeda, Y. (1999). The genes responsible for O-antigen synthesis of Vibrio cholerae O139 are closely related to those of Vibrio cholerae O22. Gene 237, 321-332.[Medline]
Zheng, J. & McIntosh, M. A. (1995). Characterization of IS1221 from Mycoplasma hyorhinis: expression of its putative transposase in Escherichia coli incorporates a ribosomal frameshift mechanism. Mol Microbiol 16, 669-685.[Medline]
Received 15 May 2000;
revised 20 June 2000;
accepted 23 June 2000.