scholarly journals An Analysis of Why Highly Similar Enzymes Evolve Differently

Genetics ◽  
2003 ◽  
Vol 163 (2) ◽  
pp. 457-466
Author(s):  
Fahd K Majiduddin ◽  
Timothy Palzkill
Keyword(s):  
Antibiotic Therapy ◽  
Amino Acid Sequence Identity ◽  
Selective Pressure ◽  
Catalytic Efficiency ◽  
Gram Negative Bacteria ◽  
Wild Type ◽  
Gram Negative ◽  
Atomic Structures ◽  
Sequence Identity ◽  
Extended Spectrum

Abstract The TEM-1 and SHV-1 β-lactamases are important contributors to resistance to β-lactam antibiotics in gram-negative bacteria. These enzymes share 68% amino acid sequence identity and their atomic structures are nearly superimposable. Extended-spectrum cephalosporins were introduced to avoid the action of these β-lactamases. The widespread use of antibiotics has led to the evolution of variant TEM and SHV enzymes that can hydrolyze extended-spectrum antibiotics. Despite being highly similar in structure, the TEM and SHV enzymes have evolved differently in response to the selective pressure of antibiotic therapy. Examples of this are at residues Arg164 and Asp179. Among TEM variants, substitutions are found only at position 164, while among SHV variants, substitutions are found only at position 179. To explain this observation, the effects of substitutions at position 164 in both TEM-1 and SHV-1 on antibiotic resistance and on enzyme catalytic efficiency were examined. Competition experiments were performed between mutants to understand why certain substitutions preferentially evolve in response to the selective pressure of antibiotic therapy. The data presented here indicate that substitutions at position Asp179 in SHV-1 and Arg164 in TEM-1 are more beneficial to bacteria because they provide increased fitness relative to either wild type or other mutants.

scholarly journals Substrate Specificity of the Escherichia coli Outer Membrane Protease OmpP

2006 ◽  
Vol 189 (2) ◽  
pp. 522-530 ◽  
Author(s):  
Bum-Yeol Hwang ◽  
Navin Varadarajan ◽  
Haixin Li ◽  
Sarah Rodriguez ◽  
Brent L. Iverson ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Amino Acid ◽  
Outer Membrane ◽  
Amino Acid Sequence Identity ◽  
Catalytic Efficiency ◽  
Wild Type ◽  
Peptide Substrate ◽  
Peptide Substrates ◽  
E Coli ◽  
Sequence Identity

ABSTRACT Escherichia coli OmpP is an F episome-encoded outer membrane protease that exhibits 71% amino acid sequence identity with OmpT. These two enzymes cleave substrate polypeptides primarily between pairs of basic amino acids. We found that, like OmpT, purified OmpP is active only in the presence of lipopolysaccharide. With optimal peptide substrates, OmpP exhibits high catalytic efficiency (k cat/Km = 3.0 × 106 M−1s−1). Analysis of the extended amino acid specificity of OmpP by substrate phage revealed that both Arg and Lys are strongly preferred at the P1 and P1′ sites of the enzyme. In addition, Thr, Arg, or Ala is preferred at P2; Leu, Ala, or Glu is preferred at P4; and Arg is preferred at P3′. Notable differences in OmpP and OmpT specificities include the greater ability of OmpP to accept Lys at the P1 or P1′, site as well as the prominence of Ser at P3 in OmpP substrates. Likewise, the OmpP P1 site could better accommodate Ser; as a result, OmpP was able to cleave a peptide substrate between Ser-Arg about 120 times more efficiently than was OmpT. Interestingly, OmpP and OmpT cleave peptides with three consecutive Arg residues at different sites, a difference in specificity that might be important in the inactivation of cationic antimicrobial peptides. Accordingly, we show that the presence of an F′ episome results in increased resistance to the antimicrobial peptide protamine both in ompT mutants and in wild-type E. coli cells.

scholarly journals Role of Autocleavage in the Function of a Type III Secretion Specificity Switch Protein in Salmonella enterica Serovar Typhimurium

mBio ◽  
2015 ◽  
Vol 6 (5) ◽  
Author(s):  
Julia V. Monjarás Feria ◽  
Matthew D. Lefebre ◽  
York-Dieter Stierhof ◽  
Jorge E. Galán ◽  
Samuel Wagner
Keyword(s):  
Host Cell ◽  
Type Iii Secretion ◽  
Effector Proteins ◽  
Switching Function ◽  
Gram Negative Bacteria ◽  
Wild Type ◽  
Gram Negative ◽  
Type Iii ◽  
Autocatalytic Cleavage ◽  
Serovar Typhimurium

ABSTRACTType III secretion systems (T3SSs) are multiprotein machines employed by many Gram-negative bacteria to inject bacterial effector proteins into eukaryotic host cells to promote bacterial survival and colonization. The core unit of T3SSs is the needle complex, a supramolecular structure that mediates the passage of the secreted proteins through the bacterial envelope. A distinct feature of the T3SS is that protein export occurs in a strictly hierarchical manner in which proteins destined to form the needle complex filament and associated structures are secreted first, followed by the secretion of effectors and the proteins that will facilitate their translocation through the target host cell membrane. The secretion hierarchy is established by complex mechanisms that involve several T3SS-associated components, including the “switch protein,” a highly conserved, inner membrane protease that undergoes autocatalytic cleavage. It has been proposed that the autocleavage of the switch protein is the trigger for substrate switching. We show here that autocleavage of theSalmonella entericaserovar Typhimurium switch protein SpaS is an unregulated process that occurs after its folding and before its incorporation into the needle complex. Needle complexes assembled with a precleaved form of SpaS function in a manner indistinguishable from that of the wild-type form. Furthermore, an engineered mutant of SpaS that is processed by an external protease also displays wild-type function. These results demonstrate that the cleavage eventper sedoes not provide a signal for substrate switching but support the hypothesis that cleavage allows the proper conformation of SpaS to render it competent for its switching function.IMPORTANCEBacterial interaction with eukaryotic hosts often involves complex molecular machines for targeted delivery of bacterial effector proteins. One such machine, the type III secretion system of some Gram-negative bacteria, serves to inject a multitude of structurally diverse bacterial proteins into the host cell. Critical to the function of these systems is their ability to secrete proteins in a strict hierarchical order, but it is unclear how the mechanism of switching works. Central to the switching mechanism is a highly conserved inner membrane protease that undergoes autocatalytic cleavage. Although it has been suggested previously that the autocleavage event is the trigger for substrate switching, we show here that this is not the case. Rather, our results show that cleavage allows the proper conformation of the protein to render it competent for its switching function. These findings may help develop inhibitors of type III secretion machines that offer novel therapeutic avenues to treat various infectious diseases.

Tolerance to glyphosate and antibiotic resistance in gram-negative bacteria isolated from soils of different agricultural management systems in Ceará, Brazil

Gaia Scientia ◽  
2021 ◽  
Vol 15 (2) ◽  
Author(s):  
Raul Vítor Ferreira de Oliveira ◽  
Margareth Borges Coutinho Gallo ◽  
Oscarina Viana de Sousa ◽  
Álef Vasconcelos Ribeiro ◽  
Tatiana Salata Lima ◽  
...  
Keyword(s):  
Stenotrophomonas Maltophilia ◽  
Selective Pressure ◽  
Multidrug Resistant ◽  
Agricultural Management ◽  
Management Systems ◽  
Cross Resistance ◽  
Gram Negative Bacteria ◽  
Conventional Farming ◽  
Gram Negative ◽  
Opportunistic Bacteria

Brazil is among the world’s largest consumers of pesticides, with glyphosate (GLY) being the most commercialized herbicide in the country. Studies showed microorganisms suffer selective pressure when exposed to pesticides, developing tolerance to pesticides and resistance to antibiotics (ABs), in a phenomenon known as “cross-resistance”. The present work aimed to evaluate the occurrence of glyphosate-tolerance and AB-resistance in bacteria isolated from different agricultural management systems in Ceará State, Brazil. Gram-negative bacteria isolated from agroforestry (S1), conventional farming (S2) and uncultivated (S3) soils were cultured in the presence of 1.6% acid glyphosate. Overall, 58 strains were isolated. Soils S1 and S2 presented several multidrug resistant (MDR) strains, the majority resistant to ampicilin. Although there was a small percentage of strains resistant to ertapenem (33%, soil S1), the fact they were found is concerning, as Carbapenem antibiotics are used to treat clinical cases of MDR bacteria, which are not common outside hospital settings. Stenotrophomonas maltophilia (soil S2), resistant to six of the eight ABs tested, was identified by MALDI-TOF mass spectrometry, and was found as one of the most common opportunistic bacteria in ICUs of Ceará hospitals.

Prevalence and Antibiogram of Extended-Spectrum Beta-Lactamase Producing Gram-negative Bacteria Isolated from Septicemic Neonates in a Tertiary Care Hospital

KYAMC Journal ◽  
2021 ◽  
Vol 11 (4) ◽  
pp. 171-175
Author(s):  
Tania Rahman ◽  
Momtaz Begum ◽  
Sharmeen Sultana ◽  
SM Shamsuzzaman
Keyword(s):  
Antimicrobial Susceptibility ◽  
Diffusion Method ◽  
Gram Negative Bacteria ◽  
Care Hospital ◽  
Beta Lactamase ◽  
Blood Samples ◽  
Gram Negative ◽  
Extended Spectrum Beta Lactamase ◽  
Extended Spectrum ◽  
Esbl Producers

Background: In recent years, Extended-spectrum beta-lactamase (ESBL) producing microorganisms have complicated treatment of infections due to resistance of ESBL producing strains to a wide range of antimicrobials. Objective: Target of this study was to determine the prevalence of ESBL producing gramnegative bacteria in neonatal sepsis cases and to reveal the antimicrobial susceptibility pattern of those isolated ESBL producers. Materials and Methods: This cross sectional study was carried out in Dhaka Medical College Hospital (DMCH) over a period of 12 months from January to December in 2016. Following isolation and identification of gram-negative bacteria from blood samples of suspected septicemic neonates, antimicrobial susceptibility test was performed by Kirby Bauer disk-diffusion method and ESBL producers were detected by Double Disk Synergy (DDS) test. Results: Among 52 Gram-negative bacteria isolated from 106 blood samples, 34.61% ESBL producers were detected and Enterobacter spp. (45%) was predominant followed by Klebsiella pneumoniae (33.33%). None of the ESBL producers was resistant to colistin and tigecycline. All ESBL producing Acinetobacter baumannii, 77.78% and 66.67% of ESBL producing Enterobacter spp and Klebsiella spp. respectively showed resistance to meropenem. All ESBL producers were resistant to piperacillintazobactam. Conclusion: Appropriate measures should be taken to prevent the spread of ESBL producing strains by combining strategies for infection prevention, control and rational use of antibiotics. KYAMC Journal Vol. 11, No.-4, January 2021, Page 171-175

scholarly journals Detection of Extended Spectrum Beta-lactamase (ESBL) Producing Gram Negative Bacteria from Clinical Specimens of Sir Salimullah Medical College and Mitford Hospital.

2017 ◽  
Vol 10 (1) ◽  
pp. 8-12
Author(s):  
Shikha Paul ◽  
Sanya Tahmina Jhora ◽  
Prashanta Prasun Dey ◽  
Bilkis Ara Begum
Keyword(s):  
Tertiary Care ◽  
Gram Negative Bacteria ◽  
Care Hospital ◽  
Beta Lactamase ◽  
Gram Negative ◽  
Clinical Specimens ◽  
Extended Spectrum Beta Lactamase ◽  
Medical College ◽  
Extended Spectrum ◽  
Esbl Producers

Detection of Extended spectrum beta lactamase (ESBL) enzyme producing bacteria in hospital settings is vital as ESBL genes are transmissible. This study was carried out to determine the distribution of ESBL producing gram negative isolates at a tertiary care hospital in Dhaka city which deals with the patients hailing from relatively low socioeconomic status.Onehundred and twenty four gram negative bacteria isolated from different clinical specimens from outpatient and inpatient departments of Sir Salimullah Medical College and Mitford Hospital (SSMC & MH) were tested for ESBL by E test ESBL method in the department of microbiology of Sir Salimullah medical college (SSMC) from March 2013 to August 2013.Out of 124 gram negative bacteria 69 (55.65%) were positive for ESBL. Among the ESBL producers, Esch.coli was the highest (46.38%) which was followed by Serratia spp (11.59%), Enterobacter spp (10.14%), Proteus spp, (8.70%), Acinetobacter spp.(7.24%) and Klebsiella spp.(5.79%). Out of 32 Esch.coli isolated from outpatient department, 10 (31.25%) were positive for ESBL. On the other hand out of 27 Esch. coli isolated from inpatient department, 22 (81.48%) were positive for ESBL. The difference was statistically significant (p<0.001).So the present study reveals that the distribution of ESBL producers is more among the hospitalized patients than the patients of the community.Bangladesh J Med Microbiol 2016; 10 (1): 8-12

scholarly journals Mining of Gram-Negative Surface-Active Enzybiotic Candidates by Sequence-Based Calculation of Physicochemical Properties

2021 ◽  
Vol 12 ◽  
Author(s):  
Roberto Vázquez ◽  
Sofía Blanco-Gañán ◽  
Susana Ruiz ◽  
Pedro García
Keyword(s):  
Physicochemical Properties ◽  
Nearest Neighbors ◽  
Regulatory Agencies ◽  
Gram Negative Bacteria ◽  
Wild Type ◽  
Gram Negative ◽  
Bacterial Surface ◽  
Negative Surface ◽  
Surface Active ◽  
Phage Lysins

Phage (endo)lysins are nowadays one of the most promising ways out of the current antibiotic resistance crisis. Either as sole therapeutics or as a complement to common antibiotic chemotherapy, lysins are already entering late clinical phases to get regulatory agencies’ authorization. Even the old paradigm of the inability of lysins to attack Gram-negative bacteria from without has already been overcome in a variety of ways: either by engineering approaches or investigating the natural mechanisms by which some wild-type lysins are able to interact with the bacterial surface. Such inherent ability of some lysins has been linked to antimicrobial peptide (AMP)-like regions, which are, on their own, a significant source for novel antimicrobials. Currently, though, many of the efforts for searching novel lysin-based antimicrobial candidates rely on experimental screenings. In this work, we have bioinformatically analyzed the C-terminal end of a collection of lysins from phages infecting the Gram-negative genus Pseudomonas. Through the computation of physicochemical properties, the probability of such regions to be an AMP was estimated by means of a predictive k-nearest neighbors (kNN) model. This way, a subset of putatively membrane-interacting lysins was obtained from the original database. Two of such candidates (named Pae87 and Ppl65) were prospectively tested in terms of muralytic, bacteriolytic, and bactericidal activity. Both of them were found to possess an activity against Pseudomonas aeruginosa and other Gram-negative bacterial pathogens, implying that the prediction of AMP-like regions could be a useful approach toward the mining of phage lysins to design and develop antimicrobials or antimicrobial parts for further engineering.

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