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Advanced Imaging & Measurement Technology Assigment

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Homework: Waters Part I — Molecular Weight

Before calculation, I visited the webpage, and copied the sequence I am working on:

eGFP sequence:

*MVSKGEELFTG VVPILVELDG DVNGHKFSVS GEGEGDATYG KLTLKFICTT GKLPVPWPTL VTTLTYGVQC FSRYPDHMKQ HDFFKSAMPE GYVQERTIFF KDDGNYKTRA EVKFEGDTLV NRIELKGIDF KEDGNILGHK LEYNYNSHNV YIMADKQKNG IKVNFKIRHN IEDGSVQLAD HYQQNTPIGD GPVLLPDNHY LSTQSALSKD PNEKRDHMVL LEFVTAAGIT LGMDELYKLE HHHHHH*

Where it contains at the end His-purification tag with (HHHHH) and a linker (LE) previously.

Then I enter to Expasy for the calculation pI/Mw: this allows estimation of the theoretical molecular weight of the protein based on its amino acid sequence, which is later used as a reference to evaluate the accuracy of the experimental mass spectrometry results.

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It determined the average: Theoretical pI/Mw: 5.90 / 28006.60

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Two adjacent charge-state peaks were selected from Figure 1 at 903.7148 and 875.4421 m/z. Using the adjacent charge-state equation,

part1equ1.png

therefore, the peak at 903.7148 m/z corresponds to charge state 31+ , and the peak at 875.4421 m/z corresponds to 32+.

The molecular weight was then calculated as:

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The experimental molecular weight shows strong agreement with the theoretical value obtained from ExPASy (28006.60 Da), indicating high measurement accuracy.

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which corresponds to 0.081% error.