نوع مقاله : مقاله پژوهشی
نویسندگان
1 دانشجوی ارشد . گروه زیست شناسی، واحد رودهن، دانشگاه آزاد اسلامی، رودهن، ایران
2 استادیار .گروه زیست شناسی، واحد رودهن، دانشگاه آزاد اسلامی، رودهن، ایران
چکیده
در تحقیق حاضر موقعیت اسیدهای آمینه آروماتیک به عنوان عناصر مهم در تشکیل و پایداری ساختار سوم آنزیم پراکسیداز تربکوهی دناتوره شده با فلز روی، با کمک آنالیز گرافهای مشتق مرتبه دوم داده های طیف سنجی مورد بررسی قرار گرفته است. برای این منظور، مشتق مرتبه دوم گرافهای جذبی و نشری با 10 تکرار در محدوده ماوراء بنفش مرئی از طیف سنجی دورنگنمایی دورانی و فلورسانس برای آنزیم در حضور 500-100 میکرومولار کلرید روی با کمک نرم افزار متلب تهیه و مطالعه شده است. نتایج نشان می دهد که یون روی فشردگی میکروساختارهای اطراف اسیدهای آمینه آروماتیک را بویژه برای تریپتوفان کاهش داده است با اینحال حتی با غلظت 500 میکرو مولار روی، تریپتوفان مستقیما در معرض حلال آبدوست قرار نمی گیرد. بطورکلی به نظر می رسد که در واسرشتگی آنزیم توسط فلز روی، تریپتوفان اهمیت بیشتری نسبت به سایر اسیدهای آمینه آروماتیک دارد.
کلیدواژهها
عنوان مقاله [English]
Evaluation of the aromatic amino acids microenvironments of zinc-denatured horseradish peroxidase; second- derivatives spectra analysis
نویسندگان [English]
- Mohammad reza Rajabi 1
- Najmeh Hadizadeh Shirazi 2
1 MSC Department of biology, Roudehen branch, Islamic Azad University, Roudehen, Iran
2 Assistant Professor, Department of biology, Roudehen branch, Islamic Azad University, Roudehen, Iran
چکیده [English]
In the present study, the position of aromatic amino acids as important elements in the formation and stability of the tertiary structure of zinc-denatured peroxidase has been investigated with the help of second-derivative graph analysis of spectroscopic data. For this purpose, the second- derivative of absorption and emission graphs with 10 replications in the visible UV range of circular dichroism and fluorescence spectroscopy for the enzyme in the presence of 500-100 μM zinc chloride was prepared and studied with the help of MATLAB software. The results show that zinc ions reduce the compaction of microstructures around aromatic amino acids, especially for tryptophan. However, even at a concentration of 500 μM zinc, tryptophan is not directly exposed to the hydrophilic solvent. In general, tryptophan seems to be more important than other aromatic amino acids in inhibiting the enzyme by zinc metal.
کلیدواژهها [English]
- Tryptophan
- Tertiary structure
- unfolding
fluorescence spectroscopy to monitor subtle changes in a monoclonal antibody structure. Journal of
Pharmaceutical Sciences,102: 52-61.
Atrooz PO, Manar A, Ibrahim A. (2016). Heavy Metals Effect on the Activity and Kinetics of Peroxidase
Enzyme in Crude Extracts of Rosmarinus officinalis and Eruca sativa. International Journal of Biochemistry
Research & Review, 15:1-8.
Chen EL, Chen YA, Chen LM, Liu ZH. (2002). Effect of copper on peroxidase activity and lignin content in
Raphanus sativus. Plant Physiology and Biochemistry, 40(5) 439-444.
Chudzik M, Macia M˛ ek-Jurczyk Z, Pawełczak B, Sułkowska A. (2016). Spectroscopic Studies on the
Molecular Ageing of Serum Albumin. Molecules, 2017: 22, 34.
Das TK, Mazumdar S. (1995). pH-Induced conformational perturbation in horseradish peroxidase. Picosecond
tryptophan fluorescence studies on native and cyanide-modified enzymes. European Journal of
Biochemistry, 227:823– 828.
Dinh NN, Winn BC, Arthur KK, Gabrielson JP. (2014).Quantitative spectral comparison by weighted spectral
difference for protein higher order structure confirmation. Analytical Biochemistry,464:60-62.
Ferrer ML., del Monte F, Mateo CR (2003) Denaturation and Leaching Study of Horseradish Peroxidase
Encapsulated in Sol-Gel Matrices. Journal of Sol-Gel Science and Technology, 26:1169–1172.
Gasymov OK, Abduragimov AR, Glasgow BJ. (2008). Site-Directed Circular Dichroism of Proteins: 1Lb
Bands of Trp Resolve Position- Specific Features in Tear Lipocalin. Analytical Biochemistry, 374:386 395.
Gasymov OK, Abduragimov AR, Glasgow BJ. (2014). Probing Tertiary Structure of Proteins Using Single
Trp Mutations with Circular Dichroism at Low Temperature. Journal of Physical Chemistry, B118: 986-
995.
Ghisaidoobe AB, Chung SJ. (2014). Intrinsic tryptophan fluorescence in the detection and analysis of proteins:
a focus on Förster resonance energy transfer techniques. International journal of molecular sciences, 15(12),
22518–22538.
Halder UC, Chakraborty J, Das N, Bose S (2012) Tryptophan dynamics in the exploration of microconformational changes of refolded β-lactoglobulin after thermal exposure: A steady state and timeresolved fluorescence approach, Journal of Photochemistry and Photobiology B: Biology, 109:50-57.
Ke Z, Ma S, Li L, Huang Q (2016) A fluorescence approach to the unfolding thermodynamics of horseradish
peroxidase based on heme degradation by hydrogen peroxide. Chemical Physics Letters, 657:49–52.
Khan AA, Rahmani AH, Aldebasi YH, Aly SM. (2014). Biochemical and pathological studies on peroxidases
-an updated review. Global journal of health science, 6(5), 87–98.
Kendrick BS, Gabrielson JP, Solsberg CW, Ma E, Wang L. (2020). Determining Spectroscopic Quantitation
Limits for Misfolded Structures. Journal of Pharmaceutical Sciences, 109:933-936.
Kumar V, Sharma VK, Kalonia DS. (2005). Second derivative tryptophan fluorescence spectroscopy as a tool
to characterize partially unfolded intermediates of proteins. International Journal of Pharmaceutics,
294:193–199.
Lasagna M, Gratton E, Jameson DM, Brunet JE. (1999). Apo horseradish Peroxidase Unfolding and Refolding:
Intrinsic Tryptophan Fluorescence Studies. Biophysical Journal, 76:443–450.
Lopez GR, Pinto DC. (2014). Silva Horseradish peroxidase (HRP) as a tool in green chemistry. RSC Advances,
4: 37244-37265.
Lucas LH, Erosy BA, Kueltzo LE, Joshi, SB, Brandau DT. (2006). Probing protein structure and dynamics by
Second - derivative ultraviolet absorption analysis of cation–p interactions. Protein Science, 15:2228–22.
Mozo-Villar´A. (2002). Second derivative fluorescence spectroscopy of tryptophan in proteins. Journal of
Biochemistry and Biophysics Methods, 50: 163–178.
Hadizadeh Shirazi N (2019). Inhibition kinetic and thermal inactivation of horseradish peroxidase in the
presence of Zn2+ ion. Journal of Food Biochemistry, e12724
Hadizadeh Shirazi N, Keihan AM, Sajjadi S. (2017). Evaluation of oxidative activity of horseradish peroxidase
in the presence of zinc ion; spectroscopic and molecular docking study. New cellular and. Molecular.
Biotechnology Journal, 27:47-54.
Nayar S, Brahma A, Mukherjee C (2002) Second derivatives fluorescence spectra of indole compounds.
Journal of Biochemistry, 131:427-435.
Ohlsson PI, Horie T, Vanderkooi JM, Paul KG (1986) Tryptophan in horseradish peroxidase. Acta Chemica
Scandinavica part B.;40(4):257-61.
Saud AM, Shahnawaz Khan M, Alhasan IM. (2019). An efficient methodology for the purification of date
palm peroxidase: Stability comparison with horseradish peroxidase (HRP). Saudi Journal of Biological
Sciences ,26(2):301-307.
Sáez-Jiménez V, Rencoret J, Rodríguez-Carvajal MA. (2016). Role of surface tryptophan for peroxidase
oxidation of nonphenolic lignin. Biotechnology and Biofuels, 9:198-203.
Sanchez KM, Gable JE, Schlamadinger DE, Kim JE. (2008). Effects of tryptophan microenvironment, soluble
domain, and vesicle size on the thermodynamics of membrane protein folding: lessons from the
transmembrane protein OmpA. Biochemistry, 47(48):12844-12852.
Strickland EH. (1974). Aromatic Contributions to Circular Dichroism Spectra of Proteins. Critical Reviews in
Biochemistry, 2: 113-175.
Sun F, Zong W, Liu R, Chai J, Liu Y. (2010). Micro environmental influences on the fluorescence of
tryptophan. Spectrochimica Acta A: Molecular and Biomolecular Spectroscopy, 76:142–145.
Thomas L, Poulos L. (2010). Thirty years of heme peroxidase structural biology. Archives of Biochemistry
and Biophysics, 500: 3–12.
Vivian JT, Callis PR (2001) Mechanisms of Tryptophan Fluorescence Shifts in Proteins, Biophysical Journal,
80(5): 2093-2109.
Vlasits J, Jakopitsch Ch, Bernroitner M, Zamocky M, Furtmüller P G. (2010). Heme peroxidases, mechanisms
of catalase activity of heme peroxidases. Archives of Biochemistry and Biophysics, 500:74–81.
Welinder K .(1979) .Amino Acid Sequence Studies of Horseradish Peroxidase Amino and Carboxyl Termini,
Cyanogen Bromide and Tryptic Fragments, the Complete Sequence, and Some Structural Characteristics
of Horseradish Peroxidase C. European Journal of Biochemistry, Y6:483-502