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3.4 Folding/Analysis β Test 1
Q1. Match the colour test with what it detects:β i-2, ii-1, iii-4, iv-3
Q2. Which techniques are used for determining the 3-D structure of proteins?β Cryo-EM, NMR and X-ray diffraction
Q3. Four peptides are at 1 mM. Which has the highest A280? (A) Ser-Val-Trp-Asp-Phe-Gly-Tyr-Trp-Alaβ Peptide A (two Trp + one Tyr)
Q4. Thorium-230 decays to radium-226 by what process?β Alpha emission
Q5. A guanosine solution has Ξ΅βββ
= 8400 Mβ»ΒΉcmβ»ΒΉ, A275 = 0.70, path 1 cm. Its concentration is:β 8.33Γ10β»Β² mmol/L
Q6. 5-HPETE shows absorbance 3.0 at 235 nm, path 1 cm, Ξ΅ = 30000 Mβ»ΒΉcmβ»ΒΉ. Its concentration is:β 100 Β΅M
Q7. Tyrosine Ξ΅ββ
β = 200 Mβ»ΒΉcmβ»ΒΉ. At what concentration (g/L) does it give O.D. 1.0 in a 0.5 cm path? (MW Tyr β 181)β 3.30
Q8. The BeerβLambert law governs:β Spectrophotometry
Q9. Carbon-14 undergoes beta decay. This causes the:β Atomic number to increase
Q10. Circular dichroism (CD) spectroscopy is used to analyse the:β Secondary structure
Q11. The helix content of a protein can be determined using a:β Circular dichroism spectrometer
Q12. The helix content of a protein can be determined using a (repeat):β Circular dichroism spectrometer
Q13. The applied centrifugal field at 5 cm from the axis at angular velocity 3000 rad/s is about:β 4.5Γ10β· cm/sΒ²
Q14. The applied centrifugal field at 5 cm from the axis at 3000 rad/s is:β 4.5Γ10β· cm/sΒ²
Q15. Which region of the CD spectrum gives details of a protein's secondary structure?β Far-UV region
Q16. Which techniques can study conformational changes in myoglobin? P.Mass spectrometry Q.Fluorescence R.Circular dichroismβ Q and R
Q17. The three-dimensional structure of a protein can be determined by:β Cryo-electron microscopy
Q18. Which amino acid does NOT contribute to intrinsic protein fluorescence?β Cysteine
Q19. How many microlitres of 20% SDS are needed to make 40 mL of 0.5% SDS?β 1 mL
Q20. What volume of an 8Γ stock buffer is needed to make 640 mL of 0.5 M (1Γ) buffer?β 80 mL
Q21. The most suitable mode to study protein conformational changes by fluorescence is:β Intrinsic fluorescence
Q22. Which one of the following is NOT a fluorophore?β Acrylamide
Q23. Which amino acid is NOT an intrinsic fluorophore of proteins?β Valine
Q24. Which would contribute to intrinsic fluorescence in a protein?β Aromatic amino acids
3.4 Folding/Analysis β Test 2
Q25. In IR spectra of proteins, the band between 1600 and 1690 cmβ»ΒΉ (amide I) corresponds to:β C=O stretching
Q26. Which isotope is used in liquid scintillation counting?β ΒΉβ΄C
Q27. Compound Z: 1.0Γ10β»β΄ M, A = 0.15, path 1 cm. Its molar extinction coefficient is:β 1500
Q28. A 1 mL LDH assay contains 10 Β΅M sodium lactate. The quantity of lactate is:β 10 nmol
Q29. The molar concentration of NaCl in a 0.876% saline (MW 58.4) is:β 0.150 M
Q30. Which statement about concentration units is correct?β Molality does not change with temperature
Q31. How many moles of NaCl are in 50 mL of a 0.15 M solution?β 0.0075
Q32. The mole fraction of glucose in a 2.315 mol/L aqueous solution is approximately:β 0.04
Q33. If 25 g NaCl is dissolved to a final 500 mL, the % (w/v) is:β 5%
Q34. Which isotope is used in PET scanning?β An iodine isotope
Q35. Phenol red has a pH transition range of:β 6.4 to 8.2
Q36. Electromagnetic radiation comes in packets called:β Photons
Q37. The protein absorption band at 210β220 nm corresponds to which electronic transition?β ΟβΟ* of peptide bonds
Q38. ΒΉΒ³ΒΉI has a half-life of 8 days. How much of a 0.5 g sample remains after 32 days?β 31.25 mg
Q39. A 2 mCi ΒΉΒ²β΅I-thyroxine sample (tΒ½ = 60 days) has what activity after 4 half-lives?β 0.125 mCi
Q40. The secondary conformation of a globular protein in solution is best determined by:β Circular dichroism
Q41. Which method CANNOT determine the secondary-structure content of a protein?β Analytical ultracentrifugation
Q42. The sedimentation velocity of a protein in a centrifuge does NOT depend on:β Charge on the protein
Q43. The sedimentation velocity of a protein depends on which property?β Shape of the protein
Q44. Tropomyosin (93 kDa) sediments at 2.6S while haemoglobin (65 kDa) sediments at 4.3S. The slow sedimentation of tropomyosin is because:β Tropomyosin is rod-shaped while haemoglobin is spherical
Q45. The sedimentation coefficient increases with:β Increase in particle mass
Q46. Heavier proteins with higher S sediment faster, yet tropomyosin (93 kDa) sediments slower than smaller proteins because it is:β Rod-shaped (high friction)
Q47. Sedimentation velocity in a centrifugal field depends on which properties?β Shape, molecular mass and density
3.4 Folding/Analysis β Test 3
Q48. To prepare a 10% solution from a 90% solution and then serial dilutions of 8, 4 and 2%, you use:β CβVβ = CβVβ dilutions with a constant final volume
Q49. Which amino acid does NOT contribute to the fluorescence of a protein?β Serine
Q50. 'Sucrose density gradient' centrifugation is used:β To purify and separate macromolecules and organelles by size/density
Q51. Tryptophan can be best identified by:β UV spectroscopy
Q52. Which method most readily identifies tryptophan?β Ultraviolet spectroscopy
Q53. When a protein is denatured by heating, its UV absorbance will:β Increase or decrease depending on the protein
Q54. The precise arrangement of atoms in a protein is determined by:β X-ray crystallography
Q55. A common method for determining the 3-D structure of proteins is:β X-ray crystallography
Q56. In X-ray crystallography, structure determination requires analysis of the:β Diffraction pattern of a protein crystal
Q57. Which centrifugation technique separates mitochondria from other organelles in a cell extract by successive speeds?β Differential centrifugation
Q58. Repeated centrifugation steps at progressively higher speeds is called:β Differential centrifugation
Q59. A common method for separating cell organelles and insoluble material from soluble proteins is:β Centrifugation
Q60. A lipid mixture on a silica column washed with increasingly polar solvents elutes in which order?β Least polar (most nonpolar) lipids elute first
Q61. Which is NOT a soft-ionization method in mass spectrometry?β Electron impact
Q62. In SDS-PAGE, breakdown of disulphide linkages is achieved by:β Ξ²-Mercaptoethanol
Q63. Haemoglobins, myoglobins and carotenoids are known as:β Chromoproteins
Q64. How many fragments does CNBr produce from Gly-Ala-His-Arg-Asp-Met-Cys-Lys-Val-Leu?β 2
Q65. How many peptides does CNBr release from Ser-Tyr-Ser-Met-Glu-His-Phe-Ser?β 2
Q66. The secondary conformation content in solution is best probed by which technique combination for myoglobin? (CD for structure)β Circular dichroism
Q67. NMR spectroscopy of biopolymers is not limited to protons. Which nucleus is also commonly observed?β ΒΉΒ³C
Q68. An advantage of NMR over X-ray crystallography is:β All of these
Q69. In an NMR spectrum, the line width is a measure of:β Relative mobility of a nucleus
Q70. An advantage of NMR spectroscopy over X-ray crystallography is that it:β Studies macromolecules in solution and reveals dynamics without crystallisation
Q71. The three-dimensional structure of a protein consisting of Ξ±-helices and Ξ²-strands can be determined by:β Nuclear magnetic resonance (NMR)
3.4 Folding/Analysis β Test 4
Q72. Which technique is used to determine the molecular weight of a protein? (NOT applicable: which one CANNOT?)β Circular dichroism
Q73. The Bradford reagent for protein estimation primarily binds to:β Basic and aromatic residues (e.g. Arg)
Q74. Arg (R), Phe (F), His (H) on cation exchange at neutral pH elute (with rising salt) in the order:β F, H, R
Q75. In cation-exchange chromatography, the solid support should be:β Negatively charged
Q76. Aspartic acid is substituted by leucine in a protein. On electrophoresis at pH 8.0, the mutant's mobility (toward the anode) will be:β Slower
Q77. Match the chromatography technique with its matrix:β I-1, II-2, III-3, IV-4
Q78. Match the stationary phase with its technique:β i-3, ii-1, iii-4, iv-2
Q79. Chromatography separates components of a mixture based on:β Differences in their distribution between stationary and mobile phases
Q80. The resolution of a chromatographic separation depends on:β The column matrix and conditions
Q81. If the dissociation constant for soluteβadsorbent binding is K_D, the retention time:β Decreases with K_D
Q82. A stationary phase for cation-exchange chromatography can be:β CM-cellulose
Q83. Two proteins P and Q catalyse the same reaction and co-localise to mitochondria. To separate them, you'd exploit their differences using chromatography based on:β Charge or size differences
Q84. Coomassie Blue staining of proteins is due to the dye interacting with:β Positively charged (basic) groups of proteins
Q85. The principle behind Coomassie Blue dye binding to protein is:β The dye's anionic groups bind protein cationic groups
Q86. On a DEAE-cellulose column, which protein appears in the flow-through: 5-lipoxygenase (pI 4.6) or patatin (pI 7.8), at the loading pH where the column binds anions?β Patatin
Q87. The binding capacity of DEAE-cellulose can be increased by equilibrating the column with:β Lower buffer ionic strength
Q88. Which method analyses protein post-translational modifications involving carbohydrate epitopes?β Eastern blotting
Q89. The separation of charged molecules under an electric current is called:β Electrophoresis
Q90. Hexapeptide P has pI 6.9. Variant Q has valine instead of glutamate at position 3. At pH 8.0, which is correct?β Q migrates slower toward the anode than P
Q91. Glu (pI 3.2), Arg (pI 10.8), Val (pI 6.0) are electrophoresed at pH 6.0. Which migrates toward the anode (+)?β Glutamic acid
Q92. Which technique determines the size of a native protein?β Gel filtration (size-exclusion) chromatography
Q93. Gel filtration chromatography separates molecules on the basis of:β Size
Q94. Which amino acid is used (in the running buffer) in SDS-PAGE?β Glycine
Q95. For detecting glycoproteins, the most commonly used probe is:β Lectins
3.4 Folding/Analysis β Test 5
Q96. Which ligand purifies a GST-tagged fusion protein by affinity chromatography?β Glutathione
Q97. Protein separation techniques exploit hydrodynamic property differences EXCEPT:β Charge of the protein
Q98. A hydrophobic protein can be purified using which chromatographic material?β Phenyl-sepharose
Q99. From a Lys/Leu/Glu mixture, which elutes FIRST from a cation-exchange resin at pH 1?β Glutamic acid
Q100. Three similarly sized peptides P(+), Q(weakly β), R(strongly β) on an ANION-exchange column elute in the order:β P, Q, R
Q101. The technique that separates based on net charge is:β Ion-exchange chromatography
Q102. Match the ion-exchanger type with its functional group:β I-4, II-3, III-1, IV-2
Q103. To separate the peptide GLEKSVRLGDVQPSLGKESRAKKFQRQ (basic, +) at pH 7.0, the appropriate matrix is a:β Carboxymethyl (cation)
Q104. Which statement about ion-exchange chromatography is NOT true?β Binding is by hydrophobic interaction
Q105. Isoelectric focusing of proteins is associated with which technique?β Electrophoresis
Q106. At pH 6, polypeptides with pI 6 will:β Not migrate (they are at their pI)
Q107. A student converts a protein's glutamic acid to glutamine (protein AβB). How can the two be resolved?β By their charge difference (e.g. ion-exchange or IEF)
Q108. Electrophoresis at pH 6 of polypeptides (mw 100, 200, 400) all with pI 6 gives:β No migration of any polypeptide
Q109. An amino acid at its isoelectric point, placed in an electric field, will:β Not move (no net charge)
Q110. Which reagent enhances colour development in Lowry's protein assay?β FolinβCiocalteu reagent
Q111. Which technique CANNOT be used to determine a protein's molecular mass?β Isoelectric focusing
Q112. Which techniques can purify a protein in its native state?β Gel filtration
Q113. Which technique can purify a protein in the native state?β Gel filtration
Q114. Which statement regarding chaperones is FALSE?β Chaperones encode the final 3-D structure
Q115. Chaperones primarily function in:β Assisting (re)folding of proteins
Q116. Which is NOT a function of molecular chaperones in protein folding?β Determining the amino acid sequence
Q117. The protein required for assisting protein folding is:β Chaperone
Q118. Specialized proteins that allow polypeptide chains to fold into stable conformations are:β Chaperones
Q119. Disulphide bonds that act as 'atomic staples' to reinforce protein conformation are formed in the:β Endoplasmic reticulum
3.4 Folding/Analysis β Test 6
Q120. Heat-shock proteins, identified as stress-induced proteins, also function as:β Molecular chaperones
Q121. Which heat-shock protein has both protein-folding and degradation-promoting activities?β Hsp100
Q122. Which force is most favourable for driving protein folding?β Hydrophobic interactions
Q123. The paradox related to protein folding is the:β Levinthal's paradox
Q124. The 'molten globule state' is associated with:β Protein folding
Q125. Which disease is associated with protein misfolding?β All of these have a misfolding component
Q126. Tryptophan fluorescence vs pH stays constant at low pH then changes β indicating that fluorescence reports on:β The local environment/conformation around Trp
Q127. The disease-causing prion PrPSc differs from normal PrPC mainly in its:β Conformation (more Ξ²-sheet)
Q128. Which method purifies an enzyme that uses arabinose as substrate from a cellular mixture?β Affinity chromatography (arabinose ligand)
Q129. Which compound specifically cleaves peptide bonds on the C-side of methionine?β Cyanogen bromide
Q130. During sequencing, the peptide bond contributed by a methionine carbonyl is cleaved by:β Cyanogen bromide
Q131. In 2-D PAGE, the second electrophoresis is run at what angle relative to the first?β 90Β°
Q132. The best chromatographic method to separate a protein that binds strongly to its substrate is:β Affinity
Q133. Which chromatographic method is best to separate a protein that binds strongly to its substrate?β Affinity chromatography
Q134. The chromatography in which the ligand is immobilized on the column matrix is:β Affinity
Q135. If a protein won't bind the matrix in affinity chromatography, which should you NOT do to troubleshoot?β Boil the protein in SDS
Q136. Match group-specific ligands: Avidinβbiotin enzymes; Proteins A/Gβimmunoglobulins; Phenylboronateβglycoproteins; LysineβrRNA/plasminogenβ Avidinβbiotin; A/GβIg; phenylboronateβglycoproteins; lysineβplasminogen
Q137. A protein precipitated above 80% ammonium sulphate saturation is likely:β Highly hydrophilic
Q138. When proteins are precipitated with ammonium sulphate, a hydrophobic protein precipitates at:β Low % saturation
Q139. Why does ammonium sulphate precipitate proteins?β It renders proteins insoluble by competing for water (salting out)
Q140. In anion-exchange chromatography, the buffer pH relative to the protein's pI must be:β Above the pI (protein negative)
Q141. In SDS-PAGE, ammonium persulphate acts as a:β Source of free radicals
Q142. In SDS-PAGE, the role of Ξ²-mercaptoethanol is to:β Cleave disulphide bonds and help denature proteins
Q143. Which method CANNOT be used to determine the molecular weight of an enzyme?β Enzyme activity assay
3.4 Folding/Analysis β Test 7
Q144. A 70 kDa heterodimer shows two SDS-PAGE bands (44 and 26 kDa). On a Sephadex G-100 (native) column it would appear as:β 70 kDa
Q145. In SDS-PAGE, ammonium persulphate provides free radicals to:β Initiate acrylamide polymerisation
Q146. In two-dimensional PAGE, the first dimension separates proteins by:β Isoelectric point (IEF)
Q147. In SDS-PAGE, one SDS molecule binds approximately how many amino acid residues?β Two
Q148. SDS is used in PAGE because it:β Solubilises and gives proteins a uniform negative charge proportional to mass
Q149. Which statement is NOT correct for SDS-PAGE components?β SDS is a cationic detergent
Q150. During SDS-PAGE, which dye is used as the tracking dye?β Bromophenol blue
Q151. In SDS-PAGE, a protein migrates in the gel until its:β Migration is determined by size, not pI (SDS masks charge)
Q152. A protein of two identical 35 kDa chains, on SDS-PAGE, migrates as:β 35 kDa
Q153. A 60 kDa homodimer is boiled with mercaptoethanol and run on SDS-PAGE. The result is:β One band at 30 kDa
Q154. SDS-PAGE separates proteins based on:β Molecular weight
Q155. In SDS-PAGE, the roles of Ξ²-mercaptoethanol and bromophenol blue respectively are:β Reducing disulphides; tracking dye
Q156. When compared with the unphosphorylated form on SDS-PAGE, a phosphorylated protein usually migrates:β Slower
Q157. Proteins A, B, C (63, 28, 79 kDa) are run on reducing SDS-PAGE. Their order from top to bottom of the gel is:β C, A, B
Q158. In size-exclusion chromatography, molecules larger than the pore size:β Cannot enter the beads and elute first
Q159. In size-exclusion chromatography:β Small molecules enter the matrix and elute later
Q160. Two proteins of 120 kDa and 25 kDa are most easily separated by:β Size-exclusion chromatography
Q161. Proteins 1 (58), 2 (28), 3 (17), 4 (10 kDa); 3 and 4 form a heterodimer, 1 and 2 are monomers. SEC elution order is:β 3+4 dimer (27 kDa) co-elutes among monomers by its combined size
Q162. In SEC of cytochrome c (13 kDa), IgG (145 kDa), RNA polymerase (450 kDa), serum albumin (68.5 kDa), the SECOND to elute is:β Immunoglobulin G
Q163. In SEC, the protein that elutes second (largest first) among the four is:β The second-largest
Q164. Proteins P, Q, R, S (50, 80, 120, 150 kDa) on Sephadex G-200 elute in the order:β S, R, Q, P
Q165. Which statement is true about size-exclusion chromatography?β Largest protein elutes first
Q166. A protein band-shift on native gel when mixing Protein M (20 kDa) and Protein X (15 kDa), with M retarded, most likely indicates:β M and X interact (form a complex)
Q167. Two proteins with the same molecular weight AND same pI are best resolved by:β Affinity chromatography (if one binds a specific ligand)
3.4 Folding/Analysis β Test 8
Q168. To separate Protein A (50 kDa, pI 5.0) and Protein B (57 kDa, pI 8.0) to high purity, the best single technique is:β Ion-exchange chromatography
Q169. Four batches of alkaline phosphatase: the most efficiently purified batch has the highest:β Specific activity
Q170. In an enzyme purification table, 'fold purification' for each step is calculated as:β Step specific activity Γ· initial specific activity
Q171. An enzyme purified 1500-fold from 70% ammonium sulphate precipitate is now pure. What fraction of total cell protein was it?β About 0.07%
Q172. In a purification table, percent yield at each step is calculated as:β (Total activity at step Γ· initial total activity) Γ 100
Q173. 'Salting in' is a process that:β Solubilises protein (increases solubility at low salt)
Q174. 'Salting in' is a process to:β Solubilise the protein
Q175. Which is considered a 'polishing step' in protein purification?β Size-exclusion (gel filtration) chromatography
Q176. For a sample labelled 10 Β΅g/mL protein, the most sensitive reagent to confirm protein is:β Lowry
Q177. 10 Β΅L protein β 990 Β΅L buffer; then 50 Β΅L of that β 450 Β΅L buffer; A = 0.1, Ξ΅ = 1.0 (mg/mL)β»ΒΉcmβ»ΒΉ, 1 cm. Starting concentration:β 10 mg/mL
Q178. Same dilutions, A = 0.1, Ξ΅ = 1.0 (mg/mL)β»ΒΉcmβ»ΒΉ: the measured (final) concentration is:β 0.1 mg/mL
Q179. A protein gives A280 = 0.34; its A(1%) (1% solution, 1 cm) is 22.9. The concentration is:β 14.9 mg/100 mL
Q180. Which methods can estimate protein concentration?β Biuret, Bradford and Lowry
Q181. Which technique determines the stoichiometry of interacting proteins?β Analytical methods like AUC/ITC give stoichiometry
Q182. Transient proteinβprotein interactions are best studied by:β Chemical cross-linking
Q183. Gelatin-PAGE (zymography) is used to identify inhibitory activity against trypsin involving which macromolecule?β Protein
Q184. In a zymographic assay for protease inhibitors, which statement is correct?β Gelatin (substrate) is co-polymerised into the gel before PAGE
Q185. A 'proteotypic peptide' is best described as:β A peptide that uniquely identifies a specific protein in MS
Q186. The radioactive molecule useful for labelling and analysing proteins is:β Β³β΅S-methionine
Q187. The half-life of the radioisotope Β³β΅S is about:β 87 days
Q188. A 5Γ10β»β΄ M tyrosine solution gives A280 = 0.75 in a 1 cm cuvette. The molar absorption coefficient is:β 1500 Mβ»ΒΉcmβ»ΒΉ
Q189. The amino acid showing characteristic absorbance at 280 nm is:β Tryptophan
Q190. A protein migrates anomalously slowly on SDS-PAGE relative to its true mass. A likely cause is:β Incomplete denaturation, glycosylation, or high proline content
3.4 Folding/Analysis β Test 9
Q191. Which is NOT involved in Edman degradation?β Dinitrofluorobenzene
Q192. Which is NOT involved in Edman degradation (repeat)?β Dinitrofluorobenzene
Q193. Which method identifies the N-terminus of a polypeptide?β Edman degradation
Q194. Which reagent is useful for determining the N-terminal amino acid?β Phenylisothiocyanate (Edman's reagent)
Q195. Which agent is commonly used for determining the N-terminal amino acid?β Phenylisothiocyanate
Q196. The first protein to be sequenced was:β Insulin
Q197. The first protein sequenced was:β Insulin
Q198. The isoelectric point is the pH at which a molecule has:β No net charge
Q199. The ninhydrin test is given by:β Proteins/amino acids
Q200. Hexapeptide composition 2R, A, V, S, Y. Trypsin: (R,A,V)+(R,S,Y); Chymotrypsin: (A,R,V,Y)+(R,S); Carboxypeptidase A: no product. The sequence is:β AVRYSR
Q201. The peptide GARAGE cleaved into two tripeptides (GAR + AGE) was cut by which protease?β Trypsin
Q202. Aminopeptidase removes residues from the N-terminus; carboxypeptidase removes from the C-terminus. These enzymes are:β Exopeptidases
Q203. Match the cleavage reagent with the residue it acts on:β I-3, II-1, III-4, IV-2
Q204. Digesting Asp-Ala-Gly-Arg-His-Cys-Asp-Lys-Trp-Lys-Pro-Ser-Glu-Asn-Leu-Ile-Arg-Thr-Tyr-Glu with trypsin gives how many peptides?β Five
Q205. Trypsin digestion of Gly-His-Phe-Leu-Arg-Ala-Gly-Met-Lys-Gly-Val-Leu gives which fragments?β GHFLR, AGMK, GVL
Q206. Trypsin cleavage of Met-Ala-Tyr-Met-Phe-Arg-Gly-Asp-Lys-Glu-Trp gives:β Met-Ala-Tyr-Met-Phe-Arg; Gly-Asp-Lys; Glu-Trp
Q207. The correct basic order for protein engineering is: I.Design II.Synthesise gene III.Express IV.Characteriseβ I, II, III, IV
Q208. Which technique separates dCTP from a mixture of dCMP, dCDP and dCTP?β Anion exchange chromatography
Q209. Which chromatography best separates dCTP from dCDP and dCMP?β Anion exchange chromatography
Q210. Which chromatographic technique uses gravity or capillary action to move the mobile phase?β Paper/thin-layer chromatography
Q211. Choose the appropriate ion-exchange matrix to bind a basic peptide (Lys/Arg-rich) at pH 7.0:β Carboxymethyl (cation exchanger)
Q212. A protein with two identical 35 kDa chains held by disulphides, on NON-reducing SDS-PAGE, runs at:β 70 kDa
Q213. Two proteins A (50 kDa) and B (~50 kDa) with the same size but different pI are best separated by:β Ion-exchange chromatography
Q214. Isoelectric focusing separates proteins on the basis of:β Isoelectric point (pI)