17 Questions 28 Answers 0 Followers
Questions related from Gustavo Avelar Molina
I am cloning five genes of approx. 1.2 kb each into pET-28a(+) vector (separately, i.e., not all in the same vector) by the USER method. The protocol is essentially the same for all constructs,...
18 February 2023 5,109 8 View
Hi all, I am thinking about other approaches than SEC to separate protein-dye conjugates from free fluorescent dyes, like Alexa, Cy and Atto dyes. Does any of you have experience with that? Will...
18 June 2021 9,291 7 View
I am looking for a reliable and fast method for separation of protein-fluorescent probe conjugates from the respective free fluorophores present in labeling reactions. HiTrap desalting columns...
07 September 2020 8,316 8 View
Dear all, I've measured the diffusion coefficient of a fluorescently labeled glycosidase interacting with its substrate (a polysaccharide) by using fluorescence correlation spectroscopy (FCS). An...
06 August 2018 5,956 4 View
Dear all, I am preparing a protocol for membrane protein reconstitution in liposomes. In this protocol, the proteins are mixed with the lipids (DLPC), and the liposomes with the proteins are...
01 November 2017 1,673 4 View
I've performed a MD simulation for a protein using GROMACS, and I want to obtain a map for the generalized correlation/dynamic cross correlation in relation to this protein. Note that I'm not sure...
10 January 2017 3,451 6 View
I have constructed a protein-carbohydrate complex based on available 3D structures. Since I did some manual adjustments in the carbohydrate structure, it is needed some kind of energy minimization...
29 October 2016 2,093 3 View
Hi everyone, Suppose one wants to elucidate the pathway of a plant secondary metabolite, or of a class of plant secondary metabolites (e.g. alkaloids) in a certain organ (e.g. fruit). How should...
28 August 2016 7,370 4 View
Dear all,When I perform FLIM-TCSPC measurements of a certain probe in solution or in a porous medium (polysaccharide substrate) I can fit the TCSPC data with only one term that corresponds to the...
29 April 2016 5,328 5 View
I have tried to separate aggregates from xylan and arabinoxylan solutions by passing them through common 0.22 um syringe filters. However, the polysaccharide content after filtration was...
20 August 2015 4,299 4 View
Most protein labeling protocols recommend to separate the free labels by extensive dialysis or by size exclusion chromatography. Does anyone know if it is possible to do the same by using Ni...
12 August 2015 9,194 5 View
Hi everyone, I am new to amylases, so I would like to know if there is any preference when it comes to substrate and assay methods choosing. Please, cite the advantages and disadvantages of each...
03 July 2015 2,709 4 View
I'm studying the binding dynamics of an enzyme to its substrate by measuring the fluorescence anisotropy of a fluorophore attached to the protein. Since this substrate is a polymer (and/or maybe...
25 February 2015 8,138 4 View
I have purified a protein by affinity chromatography (Ni Sepharose High Performance - GE Healthcare), using a 50 mM phosphate buffer pH 8.0 with 300 mM imidazole and 50 mM NaCl as an elution...
16 February 2015 3,016 14 View
I have used a PicoQuant MicroTime 200 system to do FCS measurements of free Alexa Fluor 488 dye and its conjugates with proteins in simple phosphate buffers. The samples are quite simple. However,...
11 December 2014 7,727 7 View
I need an easy way to immobilize a polysaccharide (xylan) on a cover slip glass surface for one-molecule analysis using confocal fluorescence microscopy.
08 December 2014 9,710 4 View
I want to study the interaction of this polymer with an enzyme that catalyzes its hydrolysis. To know the catalysis kinetics and the hydrolysis degree of a xylan chain in each enzyme-substrate...
27 February 2014 5,711 1 View
