Bacterial L-forms Allan, E J; Hoischen, C; Gumpert, J
Advances in applied microbiology,
2009, Letnik:
68
Journal Article
Recenzirano
L-forms are "cell wall-deficient" bacteria which are able to grow as spheroplasts or protoplasts. They can be differentiated into four types depending on their ability to revert to the parental, ...cell-walled form and to the extent of their cell-wall modification. L-forms are significant in modern science because of their contributions to an improved understanding of principal questions and their interactions with eukaryotes. This review particularly focuses on research using stable protoplast-type L-forms which have contributed to a better understanding of the structural and functional organisation of the cytoplasmic membrane and of cell division. These L-forms, which have only a single surrounding bilayer membrane, also represent a unique expression system for production of recombinant proteins. A large proportion of L-form publications concern their putative role in human disease and its therapy, a topic which is discussed briefly. L-forms have also been used to form intracellular associations with plant cells and have been shown to elicit induced disease resistance offering a novel method for plant protection. The recent decline in active research on L-forms is a concern as knowledge and experience, as well as unique L-form strains which have been maintained for decades, are being lost.
The paper describes the synthesis of the phosphorylcholine-binding miniantibody McPC603scFvDhl x in cell-wall-less L-form strains of Escherichia coli and Proteus mirabilis. Cells of these strains ...were transformed with the plasmid pACK02scKan, carrying the miniantibody (miniAb) coding sequence under the control of the lac promoter. L-form transformants of both species were able to synthesize the functional miniAb as an extracellular soluble product. The highest quantities were obtained by P. mirabilis L-form strains after induction with 5 mM isopropyl beta-D-thiogalactopyranoside (IPTG). Yields of 45-75 mg/l total antibody protein and of 10-18 mg/l functional miniAb were estimated in the growth medium of shaking cultures 40-80 h after induction with IPTG. About 10% of the active miniAb remained cell-bound. The yields of functional miniAb could be optimized by lowering the growth temperature from 37 degrees C to 26-32 degrees C and by supplementation of the medium with 80 mM sodium fumarate. A comparison of the specific activities revealed that the P. mirabilis L-form strains have a similar synthesis capacity (2-4 mg functional miniAb/g cell dry weight) to that of the producer strain E. coli RV308. The results show that the processes of correct folding and assembling of the miniAb molecules are possible without the periplasmic compartment.
AimsLarge operable cancers have traditionally been treated surgically by mastectomy. More recently centres have investigated the use of neoadjuvant chemotherapy to allow breast-conserving surgery. ...Between 1991 and 1995, a prospective study into the response of large operable breast cancers to CMF neoadjuvant chemotherapy was performed.MethodsPatients with cancers requiring mastectomy, and with or without clinically involved non-fixed lymph nodes, were offered neoadjuvant CMF chemotherapy. Patients declining neoadjuvant treatment underwent mastectomy and appropriate axillary surgery. Clinical response was assessed after two cycles in the neoadjuvant group. Subsequent surgical or non-surgical management was planned after this.ResultsThirty-eight patients were suitable for neoadjuvant treatment. Twenty-two underwent two cycles of CMF and were then reassessed. Seventy-three per cent achieved a response three (14%) complete remission, 13 (60%) partial remission. Fifteen (68%) patients avoided mastectomy, with six (27%) requiring no surgery at all with no clinically detectable residual disease. Sixteen (42%) declined neoadjuvant chemotherapy and opted for immediate mastectomy, seven of whom accepted chemotherapy post-operatively. After 3 years» follow-up there is no statistical difference in local recurrence, distant recurrence or overall survival.ConclusionApproximately 40% of patients offered neoadjuvant chemotherapy will demand prompt surgical treatment but will consider the use of adjuvant chemotherapy post-operatively. Sixty-eight per cent of patients receiving neoadjuvant CMF will successfully avoid mastectomy.
Abstract
Cell division and cell wall synthesis are tightly linked cellular processes for bacterial growth. A protoplast-type L-form Escherichia coli, strain LW1655F+, indicated that bacteria can ...divide without assembling a cell wall. However, the molecular basis of its phenotype remained unknown. To establish a first phenotype–genotype correlation, we analyzed its dcw locus, and other genes involved in division of E. coli. The analysis revealed defective ftsQ and mraY genes, truncated by a nonsense and a frame-shift mutation, respectively. Missense mutations were determined in the ftsA and ftsW products yielding amino-acid replacements at conserved positions. FtsQ and MraY, obviously nonfunctional in the L-form, are essential for cell division and cell wall synthesis, respectively, in all bacteria with a peptidoglycan-based cell wall. LW1655F+ is able to survive their loss-of-functions. This points to compensatory mechanisms for cell division in the absence of murein sacculus formation. Hence, this L-form represents an interesting model to investigate the plasticity of cell division in E. coli, and to demonstrate how concepts fundamental for bacterial life can be bypassed.
It is unclear whether patients on oral anticoagulants (OAC) undergoing a procedure using common femoral artery access have higher adverse events when compared to patients who are not anticoagulated ...at the time of the procedure.
We retrospectively reviewed data from consecutive patients who underwent a cardiac procedure at a tertiary medical center. Patients were considered (group A) fully or partially anticoagulated if they had an international normalized ratio (INR) ≥1.6 on the day of the procedure or were on warfarin or new OAC within 48 h and 24 h of the procedure, respectively. The nonanticoagulated group (group B) had an INR <1.6 or had stopped their warfarin and new OAC >48 h and >24 h preprocedure, respectively. The index primary end point of the study was defined as the composite end point of major bleeding, vascular complications, or cardiovascular-related death during index hospitalization. The 30-day primary end point was defined as the occurrence of the index primary end point and up to 30 days postprocedure.
A total of 779 patients were included in this study. Of these patients, 27 (3.5%) patients were in group A. The index primary end point was met in 11/779 (1.4%) patients. The 30-day primary composite end point was met in 18/779 (2.3%) patients. There was no difference in the primary end point at index between group A (1/27 3.7%) and group B (10/752 1.3%;
=0.3155) and no difference in the 30-day primary composite end point between group A (2/27 7.4%) and group B (16/752 2.1%;
=0.1313). Multivariable analysis showed that a low creatinine clearance (odds ratio OR =0.56;
=0.0200) and underweight patients (<60 kg; OR =3.94;
=0.0300) were independent predictors of the 30-day primary composite end point but not oral anticoagulation (
=0.1500).
Patients on OAC did not have higher 30-day major adverse events than those who were not anticoagulated at index procedure.
The structural gene of the carboxypeptidase T (
cpt) was successfully expressed in cell wall-less L-form cells of
Proteus mirabilis. The DNA sequence encoding the PhoA leader peptide was fused with a ...truncated
cpt gene encoding the mature enzyme. The modified gene in a pUC-based kanamycin resistance vector under the control of the
lac promoter was transformed into L-form cells of
P. mirabilis. They were able to produce the recombinant CpT both as a secretory and as a cell-bound insoluble form. The co-secretory processing of the PhoA leader peptide was quite efficient. The yield of the secreted CpT was not less than 20 mg l
−1 and should be improvable.
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