1. N.M. Emanuel Institute of Biochemical Physics, RAS, Moscow, Russia sdvarf@sky.chph.ras.ru Prebiological synthesis and evolution of macromolecules Sergey Varfolomeyev

2. Origin of Life = Origin of self Proliferating Macromolecules

3. Life phenomena from point of view of polymer chemistry Polycondensation – the same reaction for all biopolymers Three-functional monomers Immense decrease of number of macromolecules comparing to the number of possible variants

4. Decrease of macromolecules number Biological systems implemented the extreme low part of possible macromolecular structures Human genome3. 10 4 genes Possible variants of protein structures (20 amino acids, 100 monomers in chain) 20 100 ~ 10 130

5. Natural (biological) polymers – products of polycondensation of three-functional monomers Amino acids - polyamides (proteins) Nucleotides - polyesters (DNA, RNA)

6. Copolymerization of three- functional monomers -H 2 O

7. All biomacromolecules (products of polycondensation) are unstable in water

8. Is it possible to have polycondensation process in water? Thermocycling

9. Thermocycle – natural global cycle ΔT - 200 - +200 - 20 - + 150 0 - + 120  Polymerization at high temperatures 120 – 200 O C (statistical number of polymers with different structures and size)  Reduction of temperature, solubilization in water condensate  Macromolecular selection of monomers by polymer matrix  Temperature increasing, polymerization, monomers on matrix (template), formation of partly complementary chain

10. Natural (biological) polymers – products of polycondensation of three-functional monomers Amino acids - polyamides (proteins) Nucleotides - polyesters (DNA, RNA)

11. " Кинетика как наука о законах и механизме развития различных процессов в природе находит в области биологии почти неограниченные перспективы для теоретических исследований и практического применения." Н.М. Эмануэль, 1966

12. Copolymerization of three- functional monomers Kinetic theory describing and explain the dramatic reduction of the number of polymer structures in systems. S.D.Varfolomeyev,Kinetic models of the prebiological evolution of macromolecules.Thermocycle as motive force of the process,Mendeleev Communications,2007, 17,7-9

13. Copolymerization of three- functional monomers

14. dNi\dt =kiNi ki=k fi -k di ki >0 автоселекция ki <0 деградация Комплексообразование мономера с полимером,как правило,стабилизирует макромолекулу против термической деструкции и гидролитического расщепления

15. Copolymerization of three- functional monomers

18. Thermocycle is a method to change absolutely improbable events (probability 10 -50 ) to absolutely certain ones (probability 1). S.Varfolomeyev,Mendeleev Comm.2007,17,7-9

19. Adsorption of monomers over polymer Formation of a complex between monomer and polymer tends to stabilize macromolecule against thermal destruction and hydrolysis Synthesis of new polymer is selective

20. Synthesis

21. BA A +H 2 O BA A Hydrolysis

22. Selection of monomers on polymer and synthesis of partly complementary chain

23. Polymerization Hydrolysis Triads of two monomers Synthesis on polymer

24. Reactions

25. Equations

27. Increased stability of certain triplets AAA BBB Accumulation of AAA and BBB, no monomer competition BBA AAB AAA AA A BBA Depletion of AAB due to monomer competition with more stable BBA

28. Selection principles (competitive advantages) Greater thermodynamical stability Greater hydrolysis resistance Velocity of catalytic processes: The most stable and «quick» wins!

29. Experiments

30. Thermogravimetric analysis (TGA) Kinetics of weight reduction (water elimination) during the L-asp polycondensation time, s weight, %

31. Unique for Russia combined mass-spectrometer ICR + ionic trap Finnigan LTQ-FT

32. World of polypeptides MS/MS approach Sequencing of products of polycondensation

33. World of polypeptides

35. Lys-Asp, basic solution Chain 5 monomers: 9 structures of 60 possible Chain 7 monomers: 11 structures of 252 possible World of polypeptides KK DK DDK KDK KKK DKDK KDKK DKDD DKDDK DKKDK KDKKK

36. All structures are closely related Complimentary structures or intermediates

37. General principles of evolution of multifunctional macromolecules  Non-statistical distribution of monomers along the polymeric chain  The primary synthesized polymers works as a template for sorption and subsequent polymerization  Selection factors (stability, catalysis, etc.) Evolution to unique structure, dramatic reduction of the number of structural variants.

38. Globular peptides and proteins Flexible polyanion RNA Rigid-linear polyanion DNA Interdependent coexistence of three worlds of pre-biopolymers + + ++ - - - - - - - - - - - - -

39. In the regime of hydrolytic degradation, system accumulates peptides and nucleic acids affine to each other and forming stable supramolecule complexes Formation of stable suprmolecule complexes is a selective factor k1k1 k2k2 k3k3 Degradation k 1, k 2 > k 3 Interdependent coexistence of three worlds of pre-biopolymers

40. Accumulation of selected limited repertoire of peptides (pre-proteins) Accumulation of selected set of complexes of peptides with RNA Transfer of structural information from RNA to peptide (and vice versa) and fixation in form of double-stranded DNA, molecular hieroglyphs Polymer-RNA complex RNADNA Interdependent coexistence of three worlds of pre-biopolymers

41. The same pattern for three main groups of biopolymers Thermocycle (phase transfer (- H 2 O)) is a driving energetic force of the process and provides thermodynamics possibility of polycondensation Autoselection of polymers composed of three-functional monomers. Polymer always makes influence on the products composition due to supramolecular interactions of monomers and polymers This drives switch from statistical to evolutionary-driven synthesis Proteins and peptides RNA DNA

42. General principles of evolution of multifunctional macromolecules (1) Non-statistical distribution of monomers along the polymeric chain (2) The primary synthesized polymers work as the template for sorption and subsequent polymerization (3) Selection (stability, catalysis, etc.) (4) Evolution to unic structure, dramatic reduction of the number of structural variants.

43. Трифункциональные мономеры (X Y, X Y) Предбиологическая эволюция биополимеров  Поликонденсация - базовый химический процесс (- H 2 O), образование амидов, пептидов, белков, фосфодиэфирной (РНК, ДНК) связи  Термоцикл (фазовый переход (- H2O)) – движущая энергитическая сила процесса – обеспечение термодинамической возможности поликонденсации

44. Предбиологическая эволюция биополимеров Автоселекция полимеров из трифункциональных мономеров Полимер ВСЕГДА оказывает влияние на состав продуктов в силу супрамолекулярного взаимодействия мономера с полимером Переход от чисто статистического к эволюционно-направленному синтезу Общие закономерности для всех трех основных классов биополимеров - Пептиды и белки О, рибонуклеиновые кислоты (  ), дезоксирибонуклеиновые кислоты ( I I )

45. Большая термодинамическая устойчивость Большая устойчивость к гидролитической деструкции Каталитические свойства Гидролиз пептидов Гидролиз полинуклеотидов Гидролиз пирофосфата Получение мономеров Поликонденсация аминокислот Поликонденсация нуклеотидов Предбиологическая эволюция биополимеров Принципы отбора (конкурентные преимущества) Побеждает наиболее стабильный и «быстрый»

46.  Подвижно – линейные - РНК  Жестко - линейные - ДНК Взаимозависимое сосуществование трех «миров» предбиополимеров Глобулярные – пептиды, белки + + ++ - - - - - - -    - - - - - -

47. Взаимозависимое сосуществование трех «миров» предбиополимеров В режиме гидролитической деградации в системе накапливаются пептиды и нуклеиновые кислоты, которые афинны друг к другу, то есть образуют устойчивые супрамолекулярные комплексы Образование стабильных супрамолекулярных комплексов как фактор отбора +    k1k1 k2k2 k3k3 деградация k 1, k 2 > k 3

48. Накапливается «отселектированный» ограниченный набор структур пептидов (предбелков) Накапливается «отселектированный» набор комплексов пептидов – РНК Перенос структурной информация от пептида к РНК, и «фиксация» этой информации в ДНК - форме Взаимозависимое сосуществование трех «миров» предбиополимеров       + Комплекс Полимер - РНК РНК Цепь ДНК Двойная Спираль ДНК

49. Молекулярные иероглифы Ферменты и активные центры ферментов Антитела Рецепторы Малые РНК

50. The great paradox of enzymes origin Very restricted number of catalytical sites structures

52. 2 Metal ions (2 Zn)\HDB Parathion hydrolase (1DPM)

53. Chymotrypsin and Streptogrisin : Primary structures are absolutely different Catalytical sites are the same

54. Organophosphate hydrolase (EC 3..1.8.1) X R 2 P ZR 3 R1R1 O P OHR1R1 O + HZR 3 H 2 O Organophosphate hydrolase O X = O, S Z = O, S and F, when R 3 is absent Reaction catalyzed by organophosphate hydrolase ( EC 3.1.8.1)

55. An important conclusion: The role of Asp from catalytic triad is to orient His, but not to serve as a proton relay 2.69Å 2.62Å 2.72Å Asp102 Ser214 Ala55 Ser195 His57 X-ray structure 2PTC

56. Переход от гидролитических реакций к синтетическим- Одни и те же каталитические структуры осуществляют гидролиз и синтез

57. Structural Unity of catalytical sites PyrophosphataseExonucleaseDNA- polymerase

58. N.M. Emanuel Institute of Biochemical Physics, RAS, Moscow, Russia sdvarf@sky.chph.ras.ru Prebiological synthesis and evolution of macromolecules S.D. Varfolomeev