Download MOLECULAR BIOCHEMISTRY II INTRODUCTORY LECTURE

MOLECULAR BIOCHEMISTRY II
INTRODUCTORY LECTURE
SYLLABUS
– AMINO ACID BIOSYNTHESIS
– ENERGY METABOLISM
OBESITY
DIABETES
ATKINS DIET
– NUCLEOTIDE METABOLISM
– DNA STRUCTURE
– DNA – PROTEIN INTERACTIONS
TRANSCRIPTION FACTORS
– DNA METHYLATION
– PHOTOSYNTHESIS
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
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
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”
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
SIX ENZYME CLASSES
OXIDOREDUCTASE
TRANSFERASE
HYDROLASE
LYASE
ISOMERASE
LIGASE
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
“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
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
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
KEY CONCEPTS IN ORGANIC CHEMISTRY
THE “SIX PILLARS”
–
–
–
–
–
–
ELECTRONEGATIVITY
POLAR COVALENT BONDING
STERIC EFFECTS
INDUCTIVE EFFECTS
RESONANCE
AROMATICITY
Mullins, J.J. “Six pillars of organic chemistry”, J. Chem. Educ. 2008,
85(1), 83-87
ELECTRONEGATIVITY
Mullins, J.J. “Six pillars of organic chemistry”, J. Chem. Educ. 2008, 85(1), 83-87
POLAR COVALENT BONDING
Mullins, J.J. “Six pillars of organic chemistry”, J. Chem. Educ. 2008, 85(1), 83-87
STERIC EFFECTS
Mullins, J.J. “Six pillars of organic chemistry”, J. Chem. Educ. 2008, 85(1), 83-87
INDUCTIVE EFFECTS
Mullins, J.J. “Six pillars of organic chemistry”, J. Chem. Educ. 2008, 85(1), 83-87
RESONANCE
Mullins, J.J. “Six pillars of organic chemistry”, J. Chem. Educ. 2008, 85(1), 83-87
AROMATICITY
Mullins, J.J. “Six pillars of organic chemistry”, J. Chem. Educ. 2008, 85(1), 83-87
SUGGESTION FOR LEARNING BIOCHEMICAL
MECHANISMS
WHENEVER POSSIBLE, TRY TO RATIONALIZE
MECHANISMS USING ONE OR MORE OF THESE
“PILLARS”
AN INTRODUCTION TO AMINO
ACID METABOLISM
NITROGEN CYCLE
– THE “FIXTATION” OF NITROGEN
THE CENTRAL ROLE OF GLUTAMATE
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
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
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
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
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
THE NITROGEN CYCLE
ATMOSPHERIC N2 IS THE ULTIMATE NITROGEN SOURCE
DENITRIFICATION
N2
NO3NITROGEN FIXATION
NITRATE
REDUCTASE
NITROGENASE
NITRITE
REDUCTASE
NH4+
NO2NITRIFICATION
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
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
Kelly A., Stanley CA. (2001). “Disorders of Glutamate Metabolism”. Mental RetardAtion and Developmental Disorders. 7:287-295.
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 αKETOACIDS
– FREELY REVERSIBLE REACTIONS
 IMPORTANT IN BOTH SYNTHETIC AND DEGRADATIVE
PATHWAYS