Explore BrainMass
Share

Identification of HEPES and MES on the basis of pH and pK

This content was STOLEN from BrainMass.com - View the original, and get the already-completed solution here!

You and a lab parnter prepare buffers to use in an experiment. Your partner prepares 200 ml 50 mM MES pH 6.0 while you prepare 200 ml of 50 mM HEPES pH 6.0. (MES and HEPES are the abbreviated names for two biological buffers often used in research labs).

HEPES has MW 238.3 and pK = 7.48
MES has MW 195 and pK = 6.1

1. How many grams of HEPES did you need to weigh out to prepare this buffer?

2. Unfortunately, both you and your partner forgot to label the beakers containing the buffers. Now that the beakers are on the lab bench next to each other, you can't tell which buffer is in which beaker. The third member of your group remembers what you learned about buffers in Lab 2. She labels one beaker "A" and the other "B". She takes 5 ml of buffer from "A", adds 1.0 ml dilute HCL to the 5 ml, mixes, and checks the pH. It is 5.9. She repeats the process with buffer from "B" but the pH of this samples is now 4.0. Which beaker (A or B) has the HEPES buffer? Explain your answer.

© BrainMass Inc. brainmass.com October 24, 2018, 9:00 pm ad1c9bdddf
https://brainmass.com/chemistry/physical-chemistry/identification-hepes-mes-basis-ph-pk-114007

Solution Preview

Here comes your answers.
You and a lab parnter prepare buffers to use in an experiment. Your partner prepares 200 ml 50 mM MES pH 6.0 while you prepare 200 ml of 50 mM HEPES pH 6.0. (MES and HEPES are the abbreviated names for two biological buffers often used in research labs).

HEPES has MW 238.3 and pK = 7.48
MES has MW 195 and pK = 6.1

1. How many grams of HEPES did you need to weigh out to prepare this buffer?

We need to add 2.383 g of HEPES to ...

Solution Summary

In case of HEPES, pK> pH, hence the reaction moves to protonation. It is a stronger acid will cause the formation of HA, the protonated form. It is not a good buffer and it cannot resist pH changes, while in case of MES, pH = pK (approx 6).

$2.19
See Also This Related BrainMass Solution

Please help with the following pre-lab questions regarding protein expression in my Molecular Biology Lab

PROTEIN EXPRESSION
Underlying molecular principles:
Explain the genetic components of the lac operon regulating the IPTG-inducible expression of T7 RNA polymerase in the TrcHisB/T7 and DH5 system

Hands-On Skills:
- Inoculate and grow a transformant DH5 cell line
- Assess the cell growth phase by absorbance at 600nm
- Induce protein expression by adding IPTG
- Harvest cells from a liquid culture
- Prepare one buffer solution to be used in a future laboratory class

I need help on the following two questions:
1. Perform a quick review of literature on recombinant protein expression in E. coli and indicate the protein yield reported in two research articles of your choice. Make sure you specify the units for the protein yield. For each article describe the nature of the protein that was overexpressed (membrane protein, cytosolic protein, ...). Estimate how much of your recombinant T7 RNA polymerase enzyme you might expect from 50ml of liquid culture by assuming a yield comparable to the ones reported in the two articles you selected. A hard copy of each of your two articles along with your answer should be handed in to your TA when entering the lab

2. 2. The overview figure provided in Experiment A shows the planning for the control aliquots to be assessed by SDS-PAGE in lab 6. Due to space limitation in the shaking incubator, some controls will be done collectively. Explain the information to be obtained from each of the two extra sets of controls to be done collectively:
a. The inoculated 50ml culture containing 200ug/ml ampicillin is to be divided in two fractions of 25ml. At time zero of the induction, the first 25ml fraction is to be induced with 1mM IPTG whereas only water is to be added to the second 25ml fraction. A 1ml aliquot is to be withdrawn from each of the two fractions at both time zero and after a 1.5 hr incubation. ( /2.5 MARKS)
b. A liquid culture of non transformed DH5 cells will be incubated without ampicillin, and a 1ml aliquot is to be collected at time zero and the very end of the induction treatment with IPTG.

Thanks again.

View Full Posting Details