SOME CHEMICAL PRINCIPLES TO BE COVERED

Slide 2:SOME CHEMICAL PRINCIPLES TO BE COVERED

BIOCHEMICAL PATHWAYS ENZYME CLASSIFICATION MECHANISMS REGULATORY CONTROL ROLE OF METAL IONS IN BIOCHEMISTRY PRINCIPLES OF CATALYSIS TRANSITION STATES COFACTORS ADDITION OF C1 UNITS OXIDATION/REDUCTION REACTIONS

Slide 3:ENZYME CLASSIFICATION

SIX CLASSES ( http://us.expasy.org/enzyme/ ) NOMENCLATURE COMMITTEE OF INTERNATIONAL UNION OF BIOCHEMISTRY AND MOLECULAR BIOLOGY (1992) COVALENT CHEMICAL BONDS MADE/BROKEN OXIDOREDUCTASES TRANSFERASES HYDROLASES LYASES ISOMERASES LIGASES ADDITIONAL CLASS (�ENERGASES�) PHYSICAL REACTIONS NON-COVALENT PRODUCT-LIKE AND SUBSTRATE-LIKE STATES

Slide 4:WHAT CONSTITUTES A CHEMICAL BOND?

��there is a chemical bond between two atoms or groups of atoms in case that the forces acting between them are such as to lead to the formation of an aggregate with sufficient stability to make it convenient for the chemist to consider it as an independent molecular species.� Linus Pauling in �The Nature of the Chemical Bond�

Slide 5:SIX TRADITIONAL ENZYME CLASSES

CAN YOU RECOGNIZE THE CLASS TO WHICH AN ENZYME BELONGS BY LOOKING AT THE OVERALL REACTION? IN-CLASS EXERCISE FOR THE FOLLOWING 10 REACTIONS WHICH YOU HAVE ALREADY SEEN THUS FAR IN YOUR STUDY OF BIOCHEMISTRY, INDICATE THE ENZYME BY NAME OR BY CLASS

Slide 6: SIX ENZYME CLASSES

OXIDOREDUCTASE TRANSFERASE HYDROLASE LYASE ISOMERASE LIGASE

Slide 7:CATALYSIS OF �PHYSICAL� REACTIONS

PRODUCT-LIKE AND SUBSTRATE-LIKE STATES: EXAMPLES : CHAPERONIN-MEDIATED (PROTEIN FOLDING) CHROMATIN CONDENSATION �MOLECULAR MOTOR� OPERATION DNA PROCESSING BY POLYMERASES ACTIVE AND CARRIER-MEDIATED TRANSPORT G-PROTEIN MEDIATED REGULATION OF HORMONE RECEPTORS MEMBRANE TRANSPORTERS (PUMPS) ARE NOW RECOGNIZED AS A SPECIAL CLASS OF ENZYMES �ENERGASES� : TRANSDUCE ENERGY FROM COVALENT BONDS INTO MECHANICAL WORK

Slide 8:�ENERGASES�

MEDIATE NUCLEOSIDE TRIPHOSPHATE HYDROLYSIS THE FREE ENERGY RELEASED IS COUPLED TO SYSTEM�S CONFORMATIONAL CHANGE ARE ATPases AND GTPases CORRECTLY CLASSIFIED AS �HYDROLASES�? ATP + H2O ? ADP + Pi + HEAT Keq = [ADP][Pi] / [ATP] ?Ghydrolysis IS RELEASED AS HEAT HERE THE ENZYME IS ATPase AND IT�S A HYDROLASE

Slide 9:ENERGASE EXAMPLE

A SYNTHETASE REACTION: ATP + GLU + NH3 ? GLN + ADP + Pi HERE THE ?Ghydrolysis IS COUPLED TO ?Gsynthesis THROUGH A REACTIVE INTERMEDIATE Keq = [GLN][ADP][Pi] / [ATP][GLU][NH3] = [GLN] / [GLU][NH3] X [ADP][Pi] / [ATP] AN ENERGASE REACTION: ATP + STATE 1 + H2O ? ADP + STATE 2 + Pi HERE THE ?Ghydrolysis IS COUPLED TO ?Gconformational change Keq = [STATE 1] / [STATE 2] X [ADP][Pi] / [ATP] NOTICE SIMILARITY TO Keq FOR SYNTHETASE REACTION THERE�S NO CHEMICAL (COVALENT) CHANGE, THOUGH

Slide 10:ENZYMES AS MECHANOCHEMICAL PROTEINS

THE GIBBS FREE ENERGY OF ATP HYDROLYSIS IS TRANSDUCED INTO A FORM OF USEFUL WORK TRANSLATION ROTATION SOLUTE GRADIENT A RECIPROCAL RELATIONSHIP ENZYMES USE NON-COVALENT INTERACTIONS TO BREAK COVALENT BONDS ENERGY FROM BREAKING COVALENT BONDS CAN MODIFY NON-COVALENT INTERACTIONS

Slide 19:AN INTRODUCTION TO AMINO ACID METABOLISM

NITROGEN CYCLE THE �FIXTATION� OF NITROGEN THE CENTRAL ROLE OF GLUTAMATE

Slide 20:THE NITROGEN CYCLE

N2 IS A VERY STABLE MOLECULE BOND ENERGY = 941.4 kJ/MOL COMPARED TO 498.7 kJ/MOL FOR O2 A SINGLE C=O BOND IN CO2 IS 799 kJ/MOL HOW IS IT METABOLIZED (�FIXED�)? THE �NITROGEN CYCLE� PRODUCTION OF METABOLICALLY USEFUL NITROGEN NITRITES NITRATES AMMONIA

Slide 21:THE NITROGEN CYCLE

N-FIXING ORGANISMS: ANAEROBES MARINE CYANOBACTERIA �DIAZOTROPHS� DIAZOTROPHS COLONIZE ROOT NODULES OF LEGUMES GENUS Rhizobium SYMBIOTIC RELATIONSHIP ENZYME IS �NITROGENASE� THE NITROGENASE REACTION: N2 + 8 H+ + 8 e- + 16 ATP + 16 H2O ? 2 NH3 + H2 + 16 ADP + 16 Pi REQUIRES ATP AND ELECTRONS CONTAINS Fe AND Mo

Slide 22:THE NITROGEN CYCLE

ENERGETICALLY COSTLY NEED 16 ATPs TO �FIX� ONE N2 MOLECULE COMPARE THIS TO INDUSTRIAL FIXATION: TEMPERATURE 300o - 500o C PRESSURE > 300 ATM METAL CATALYST NH3 FORMED IS USED IN FORMATION OF GLUTAMATE (Glu Dehydrogenase) GLUTAMINE (Gln Synthetase) EXCESS NH3 EXCRETED INTO SOIL RESTORE USABLE NITROGEN BY PLANTING ALFALFA

Slide 23:THE NITROGEN CYCLE

MOST PLANTS DO NOT SUPPORT N-FIXING BACTERIA NEED PRE-FIXED NITROGEN SOURCE NH3 NO2- NO3- SOURCES: LIGHTNING (10% OF NATURALLY-FIXED N) FERTILIZERS DECAY OF ORGANIC MATTER IN SOIL

Slide 24:THE NITROGEN CYCLE

PLANTS, FUNGI, BACTERIA REDUCE NO3-: A TWO-STEP PROCESS NO3- + 2H+ + 2e- ? NO2- + H2O ENZYME: NITRATE REDUCTASE NO2- + 8H+ + 6e- ? NH4+ + 2H2O ENZYME: NITRITE REDUCTASE SOME BACTERIA CAN OXIDIZE NH4+ �NITRIFICATION� NH4+ ? NO2- AND THEN TO NO3- DENITRIFICATION CONVERSION OF NO3- TO N2 BY OTHER BACTERIA

Slide 25:THE NITROGEN CYCLE

ATMOSPHERIC N2 IS THE ULTIMATE NITROGEN SOURCE N2 NH4+ NO3- NO2- NITROGEN FIXATION NITRIFICATION DENITRIFICATION NITROGENASE NITRATE REDUCTASE NITRITE REDUCTASE

Slide 26:ORGANISMS ASSIMILATE NH3

ROLE OF GLUTAMINE SYNTHETASE MICRO-ORGANISMS: ENTRY POINT FOR FIXED N GLU + ATP + NH4+ ? GLN + ADP + Pi IN ALL ORGANISMS, GLN IS AN AMINO GROUP CARRIER GLUTAMATE SYNTHASE IN BACTERIA, PLANTS ?-KETOGLUTARATE + GLN + NADPH + H+ ? 2 GLU + NADP+ OVERALL RXN�: ?-KG + NH4+ + ATP + NADPH + H+ ? GLU + NADP+ + ADP + Pi

Slide 27:THE CENTRAL ROLE OF GLUTAMATE

�GLUTAMATE FAMILY� OF AMINO ACIDS DEGRADATIVE METABOLISM CONVERGES ON THAT OF GLU GLU GLN PRO HIS ARG ORNITHINE GLU IS THE PRECURSOR OF PRO ORNITHINE ARG GLU/?-KG ARE TRANSAMINATION PARTNERS AMINO ACID + ?-KG ? GLU + ?-KETOACID OXIDATIVE DEAMINATION OF GLU (GLU DEHYDROGENASE) GLU + NAD(P)+ + H2O ? ?-KG + NAD(P)H + NH4+ N-ACETYLGLUTAMATE SYNTHESIS ALLOSTERICALLY REGULATES CPS I OF UREA CYCLE GLU + ACETYL-CoA ? N-ACETYL GLUTAMATE

Slide 29:CLOSING POINTS

HIGH ENERGY COSTS TO FIX NITROGEN ITS USE MUST BE CAREFULLY CONTROLLED GLU AND GLN ARE PIVOTAL IN AMINO GROUP TRANSFER GLU OFTEN DONATES THE AMINO GROUP GLN STORES, CARRIES AMINO GROUPS TRANSAMINASES CATALYSTS FOR TRANSFER OF AMINO GROUPS TO a-KETOACIDS FREELY REVERSIBLE REACTIONS ? IMPORTANT IN BOTH SYNTHETIC AND DEGRADATIVE PATHWAYS