proposed the sequense of protein

BMSE 205B Protein Characterization Final – Winter 2024 Please answer the following questions concisely. Basic advice for meritorious answers is to state your assumptions and show the key steps leading to your conclusions. The Veil of Veronica of Manopello is valued second only to the Shroud of Turin by the Vatican and remains equally as controversial. The Veil is basically a sheet of woven linen on which appears the gray-scale image of a man’s face beset by extreme physical suffering and alleged by some to be Christ and by others to be St John, but radiocarbon-dated to the medieval period. The image has stirred interest and controversy because no one understands what it is made of. In the 1980s, experts at various national forensic institutes were permitted to do chemical analyses of the fabric. Abundant cellulose in hydrolyzed fibers scraped from the edges of the Veil was expected given the linen. However, a significant residue of protein was also solubilized upon stirring 1 g of fibers for 2 days in 6M guanidine hydrochloride (pH 6.5; 10 °C). Church authorities ended all access to the Veil in 1988. The following biochemical data were collected at the time of the halt: 1. 2. 3. 4. 5. Peptide has apparent molecular weight ~2.7K (SDS PAGE) The pI of the unfolded peptide was about 10.0 (iso-electric focusing) Chymotrypsin digestion produced four peptides The C-terminus of largest chymotryptic peptide was HHVGNF (CPY ladder) The native N terminal residue was Ala by manual Edman chemistry. 6. The native C-terminus was blocked to CPY-ladder sequencing Investigators at the Swiss Sankt Gallen Institute of Textiles in Switzerland subsequently added the following analyses to 50 µg of already stockpiled crude protein: A. ESI spectrum: m/z at 1337.23, 891.82, 669.12, 535.50, 446.42 (monoisotopic) B. 5 µg hydrolyzed contains 3.75 nmoles of isoleucine C. The MALDI TOF monoisotopic m/z values for fragments obtained by chymotrypsin digestion in borate pH 8.2 were 237.1; 501.2; 489.1; 1503.2 D. Digestion with Arg-C-endopeptidase produced two fragments m/z 804.5 and 1887.96. E. 3.80 nmoles ammonia were detected following hydrolysis of 4.9 µg protein. F. MS/MS with CID for the chymotryptic fragment [M+H]+ = 501.2 (monoisotopic). a. y” and b ions: 402, 320, 279, 223, 166, 100 I. Propose a mass and sequence that is consistent with as many data as possible; II. Answer the following with the help of the databases: 1. 2. 3. 4. What protein is it? What organism is it from? What function is usually associated with the protein? What processing has the protein undergone? There is a very rational and natural way by which this protein came to be associated with cellulose. What is it? 5. The crude GuHCL extract contains >20 protein impurities. Propose a rational purification scheme for the protein that involves no more than four chromatographic steps. Due back in hard copy, Wed, Feb. 17th at noon.

proposed the sequense of protein

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