scholarly journals MALDI-TOF MS and CD Spectral Analysis for Identification and Structure Prediction of a Purified, Novel, Organic Solvent Stable, Fibrinolytic Metalloprotease fromBacillus cereusB80

2015 ◽  
Vol 2015 ◽  
pp. 1-13 ◽  
Author(s):  
Rajshree Saxena ◽  
Rajni Singh
Keyword(s):  
Bacillus Cereus ◽  
Protein Function ◽  
Structure Prediction ◽  
Maldi Tof Ms ◽  
Maldi Tof ◽  
C Terminus ◽  
Sds Page ◽  
Zinc Metalloproteases ◽  
Tof Ms

The ability to predict protein function from structure is becoming increasingly important; hence, elucidation and determination of protein structure become the major steps in proteomics. The present study was undertaken for identification of metalloprotease produced byBacillus cereusB80 and recognition of characteristics that can be industrially exploited. The enzyme was purified in three steps combining precipitation and chromatographic methods resulting in 33.5% recovery with 13.1-fold purification of enzyme which was detected as a single band with a molecular mass of 26 kDa approximately in SDS-PAGE and zymogram. The MALDI-TOF MS showed that the enzyme exhibited 70–93% similarity with zinc metalloproteases from various strainsBacillussp. specifically fromBacillus cereusgroup. The sequence alignment revealed the presence of zinc-binding region VVVHEMCHMV in the most conserved C terminus region. Secondary structure of the enzyme was obtained by CD spectra and I-TASSER. The enzyme kinetics revealed a Michaelis constant(Km)of 0.140 μmol/ml andVmaxof 2.11 μmol/min. The application studies showed that the enzyme was able to hydrolyze various proteins with highest affinity towards casein followed by BSA and gelatin. The enzyme exhibited strong fibrinolytic, collagenolytic, and gelatinolytic properties and stability in various organic solvents.

Molecular weight determination of high molecular mass (glyco)proteins using CGE-on-a-chip, planar SDS-PAGE and MALDI-TOF-MS

2010 ◽  
Vol 31 (23-24) ◽  
pp. 3850-3862 ◽  
Author(s):  
Roland Müller ◽  
Martina Marchetti-Deschmann ◽  
Helmut Elgass ◽  
Heimo Breiteneder ◽  
Martin Kratzmeier ◽  
...  
Keyword(s):  
Molecular Weight ◽  
Molecular Mass ◽  
High Molecular Mass ◽  
Molecular Weight Determination ◽  
Maldi Tof Ms ◽  
Maldi Tof ◽  
Sds Page ◽  
Tof Ms

scholarly journals Determination of Emedastine in Plasma by the MALDI-TOF MS.

2000 ◽  
Vol 48 (2) ◽  
pp. 150-154 ◽  
Author(s):  
Yoichiro NIHASHI ◽  
Mitsuhiro ISHIDA ◽  
Fumio SAKAMOTO ◽  
Yuichiro NAKADA
Keyword(s):  
Maldi Tof Ms ◽  
Maldi Tof ◽  
Tof Ms

Electroelution of Intact Proteins from SDS-PAGE Gels and Their Subsequent MALDI-TOF MS Analysis

2006 ◽  
pp. 353-364 ◽  
Author(s):  
Zhentian Lei ◽  
Ajith Anand ◽  
Kirankumar S. Mysore ◽  
Lloyd W. Sumner
Keyword(s):  
Maldi Tof Ms ◽  
Intact Proteins ◽  
Maldi Tof ◽  
Ms Analysis ◽  
Sds Page ◽  
Tof Ms

scholarly journals 2155. Accelerated Confirmation of Porin Loss in Carbapenem-Resistant Enterobacterales: A MALDI-TOF Mass Spectrometry-Based Approach

2019 ◽  
Vol 6 (Supplement_2) ◽  
pp. S731-S731
Author(s):  
Carlos Correa-Martinez ◽  
Evgeny A Idelevich ◽  
Karsten Becker
Keyword(s):  
Mass Spectrometry ◽  
Molecular Weight ◽  
Gold Standard ◽  
Sodium Dodecylsulfate ◽  
Resistant Bacteria ◽  
Maldi Tof Ms ◽  
Maldi Tof ◽  
Carbapenem Resistant ◽  
Sds Page ◽  
Tof Ms

Abstract Background The accurate identification of carbapenem resistance mechanisms is decisive for the appropriate selection of antibiotic regimens. Numerous methods can detect carbapenemase-producing carbapenem-resistant bacteria (CPCR). However, non-CPCR (NCPCR) are routinely assumed to display porin loss as a diagnosis of exclusion. No further confirmatory tests are performed since the gold standard (sodium dodecylsulfate polyacrylamide gel electrophoresis, SDS–PAGE) is laborious and time consuming. We propose a test for rapid and easy detection of porin loss by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS). Methods Clinical meropenem-resistant Enterobacterales strains (10 CPCR, 10 NCPCR) and control strains recommended by EUCAST (5 carbapenemase-producing, one with porin loss, one-negative control) were analyzed. Membrane proteins were extracted by successive centrifugation of bacterial suspensions (McFarland 0.5) and addition of ethanol, formic acid and acetonitrile. MALDI-TOF MS of the protein extracts was performed on a 96-spot target (Bruker Daltonics, Germany). Peaks between 35 and 40 kDa were analyzed for the presence of porins and compared with the bands observed in the SDS–PAGE of the protein extracts. Results Within the molecular weight range of 35–40 kDa, the MALDI-TOF MS-based method revealed peaks in all CPCR isolates corresponding to those observed in the carbapenemase-producing control strains. In contrast, the control strain with porin loss as well as all CNCR isolates showed a lower quantity of peaks in this range. All peaks observed correlated with the bands observed in the SDS–PAGE of the protein extracts at the corresponding molecular weight (Figure 1). Conclusion Yielding results that reliably correspond to the current gold standard, we propose a method for accelerated detection of porin loss as an alternative to the diagnosis of exclusion usually made in routine settings. With a processing time of approximately 20 minutes, the method can be easily implemented in the clinical setting. Applying this MALDI-TOF MS-based approach, valuable information will be provided about a resistance mechanism that otherwise remains unexplained. Disclosures All authors: No reported disclosures.

Determination of sulfonated azo dyes in chili powders by MALDI-TOF MS

2019 ◽  
Vol 411 (22) ◽  
pp. 5833-5843 ◽  
Author(s):  
Missael Antonio Arroyo Negrete ◽  
Kazimierz Wrobel ◽  
Eunice Yanez Barrientos ◽  
Alma Rosa Corrales Escobosa ◽  
Francisco Javier Acevedo Aguilar ◽  
...  
Keyword(s):  
Azo Dyes ◽  
Maldi Tof Ms ◽  
Maldi Tof ◽  
Tof Ms ◽  
Sulfonated Azo Dyes

A magnetite/oxidized carbon nanotube composite used as an adsorbent and a matrix of MALDI-TOF-MS for the determination of benzo[a]pyrene

2011 ◽  
Vol 47 (35) ◽  
pp. 9816 ◽  
Author(s):  
Xiao-Shui Li ◽  
Jian-Hong Wu ◽  
Li-Dan Xu ◽  
Qin Zhao ◽  
Yan-Bo Luo ◽  
...  
Keyword(s):  
Carbon Nanotube ◽  
Maldi Tof Ms ◽  
Maldi Tof ◽  
Oxidized Carbon ◽  
Carbon Nanotube Composite ◽  
Tof Ms
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