LIPIDS by hcj



   Typical biological membrane
    Phospholipid bilayer with proteins
    Extrinsic membrane proteins are attached to the
    Intrinsic membrane proteins are inserted into
the lipid bilayer

Three Main Classes of Membrane Proteins:

 a) Transport proteins

 b) Enzymes for later stages in biosynthesis of:
   phospholipids, lipopolysaccharide, peptidoglycan
 and other envelope components

 c) Energy transduction:

       Energy generation by respiration or
       Interconversion of PMF and ATP
       Energy consumption
Types of Lipid
a) Waxes - esters of a single long chain fatty acid with a
    long chain alcohol. Found on surfaces of plants,
    insects, etc. Not found in bacteria.

b) Fats - esters of glycerol with 3 long chain fatty acids
    (i.e. triglycerides). Found as globules in eukaryotes.
    Absent from most bacteria.

c) Complex lipids - esters of glycerol with two fatty
    acids. Third position of glycerol is attached to
    another chemical grouping which may be:
         phosphate plus base - phospholipids
         sulfate (plus sugar) - sulfolipids
         sugars - glycolipids

        Glycolipids are found in eukaryotes and some

        Sulfolipids are mostly found in photosynthetic

        Phospholipids make up lipid portion of the
        cytoplasmic membrane

        The outer membrane of gram-negative bacteria:
            inner half contains phospholipid
            outer half contains lipopolysaccharide (LPS)

        Lipid portion of LPS is called lipid A = an unusual
        type of glycolipid

d) Isoprenoids - repeating 5-carbon unit with CH3 branches
    Steroids of eukaryotes
    Side chains of quinones and chlorophylls
    Carrier lipids
    Membranes of Archaebacteria contain long isoprenoid
    chains instead of normal fatty acids
Fatty Acids in the Phospholipids of Bacteria
       Phospholipid = Glycerol + 2 fatty acids + phosphate +

    Bacteria contain mostly C16 fatty acids with some C18
    Note that they are all even-numbered – made from C2
    Either saturated or one double bond
    Traces of longer and shorter fatty acids also occur

The fatty acids of E. coli are mostly:
       Palmitic acid (16:0)       CH3 (CH2)14 COOH
       Palmitoleic acid (16:1)        CH3 (CH2)5 CH=CH(CH2)7
       cis-Vaccenic acid (18:1)   CH3 (CH2)5 CH=CH(CH2)9 COOH

Fatty Acids in Eukaryotes
Eukaryotic membranes are slightly wider than bacterial
membranes because the average chain length of the fatty
acids is longer
Eukaryotes and cyanobacteria also have fatty acids with
multiple double bonds
Eukaryotes usually have oleic acid instead of cis-vaccenate
- differ only in the position of the double bond
       Oleic acid (18:1)      CH3 (CH2)7 CH=CH(CH2)7 COOH

Membrane Fluidity
Membranes are viscous two-dimensional liquids

Proteins float in or on the lipid and may drift sideways
    Incorporation of transport proteins into a rigid lipid
bilayer can occur, but the protein is non-functional

Fatty acid chains in the lipid bilayer must be semi-fluid
    Correct ratio of saturated to unsaturated fatty acids
is needed
All cells increase the ratio of UFA to SFA as the
temperature falls
Between the peptidoglycan and the outer membrane

20 to 40% of total volume of cells grown under
typical conditions

It contains the following:

  a)   Binding proteins for transport for arabinose,
       maltose, galactose, histidine, leucine,
       several other amino acids and sugars,
       phosphate, sulfate, vitamin B12, vitamin B1

  b)   Scavenging enzymes such as asparaginase, acid
       phosphatase, alkaline phosphatase,
       carboxypeptidase-II, endonuclease-I, 5'-
       nucleotidase (UDP-glucose hydrolase)

  c)   Protective enzymes which inactivate
              e.       -lactamase

  d)   Membrane derived oligosaccharides (MDO) that
       contain 8 to 10 glucose residues with
       phosphoglycerol side-chains. These increase
       when osmolarity of the medium drops.
          Synthesis of Peptidoglycan

General principles in polysaccharide
   a) Use of nucleoside triphosphate (NTP) to activate
              sugar-1-P + NTP      -sugar + PPi

   This gives NDP-sugar (sugar-P-P-ribose-base) as
      activated precursors

   N may be any of A, T, G, C, or U depending on the

   b) Use of polyisoprenoid carrier lipids (bactoprenol-P
      in bacteria) to carry water soluble precursors
      across the membrane

Stages of Peptidoglycan Synthesis:

 1) Make N-acetyl-glucosamine (NAG) & N-acetyl-muramic
    acid (NAM)
 2) Activate sugars with UTP
 3) Add pentapeptide sidechain to UDP-NAM
 4) Attach UDP-NAM-pentapeptide to bactoprenol-P
 5) Add NAG
 6) Export subunit across cytoplasmic membrane
 7) Transfer subunit from bactoprenol to growing chain
 8) Cross link side chains
Stages (1) and (2) actually occur muddled together:

                Fructose-6-P 
                  
                   + Glutamine
                  
                  
                   + Acetyl CoA
                  
                  
                  
                  
                   + UTP
                  
                  
                   + PEP + NADPH
                  

Stage (3): Add Pentapeptide Side-chain

Add 5 amino acids to –COOH on 3-carbon side-chain of NAM
      In E. coli these are: L-Ala, D-Glu, meso-DAP, D-Ala-D-

The final addition is a pre-made dipeptide of two D-
      a)   Racemase:     L-Alanine  D-Alanine
      b)   Synthetase:   D-Ala + D-Ala  D-Ala-D-Ala
We now have: UDP-NAG and UDP-NAM-pentapeptide

So far reactions are in the cytoplasm – we must next cross
the membrane

Stages (4) and (5): Bactoprenol Carrier

Bactoprenol carries precursors across the inner membrane

Bactoprenol is a polyisoprenoid
    Also called undecaprenol (11 isoprene units of 5
    carbons each)
    In eukaryotes carrier = dolichol-phosphate (twenty
    isoprene units)

        + UDP-NAM-pentapeptide
        + UDP-NAG

Stage (6) Chain Elongation

NAG/NAM-pentapeptide is transferred from bactoprenol to
growing chain

Bactoprenol-PP is released and is converted back to

Stage (7) Cross Linking
Antibiotics Which Affect Cell Wall Synthesis


Prevents formation of UDP-muramic acid
       PEP analog - inhibits reaction of PEP with UDP-NAG

No practical importance - bacteria mutate to resistance
very easily by losing transport system
                               H         P         CH3               P
            phosphoenol            C C                   C       C
            pyruvate                                                       phosphonomycin
                               H         COOH        H       O       H


                                   CH3                   O
                                             NH2                     NH2
                D-alanine       HO                       N
                                                    H                      D-cycloserine
                                         O                       O

Analog of D-Alanine
       Inhibits both D-Ala racemase and D-Ala-D-Ala synthetase

Binds 100-fold better to these enzymes than D-Ala
       Rigid ring-structure of D-cycloserine holds it in one

Has side effects on synthesis of brain lipids

Note that bacitracin contains D-amino acids in its
                 S              CH2 CH3
                      CH CH
                                CH3             DOrn   Ile
                 N    NH2
          O C
                Leu   DGlu          Ile   Lys                DPhe

                       bacitracin               Asp    His

Binds bactoprenol-PP and stops conversion back to

Stops synthesis of peptidoglycan, O-antigen of LPS, capsule

Outer membrane of gram-negatives protects them against


Glycopeptide made of sugars and amino acids

MW = 1500 - does not cross outer membrane of gram-negatives

Used against methicillin resistant Staph (“MRSA”) and Strep

Vancomycin binds to D-Ala-D-Ala and blocks cross linking

Two mechanisms of vancomycin resistance.
    I) Thicker peptidoglycan
    II) Modify ends of pentapeptide from D-Ala-D-Ala to D-
         Vancomycin no longer binds
Beta-Lactam Antibiotics: Penicillins
   and Cephalosporins
Beta-lactam ring fused to a second ring that contains
Lactam = a cyclic amide;                beta-lactam has ring size = 4
     Second ring is 5-membered in penicillins
                        6-membered in cephalosporins

               penicillin nucleus              ce phalosporin nucleus
                                                         H H
                        H H       CH3                           S
                              S              R1 CONH
                                    CH3                     N
                          N                          O                  R2
                    O               COOH

Molds make the nucleus
Different molds make different beta-lactams
Side chain (R) from chemicals added to fermenting mold
     If phenylacetic acid is added the product is benzyl-
penicillin (Pen G)

Beta-lactams only affect cells which are actively growing

     Pre-made peptidoglycan is not affected

Need 1000x as much to kill gram negatives as gram positives
     due to exclusion by the outer membrane

Cells are killed if external medium is hypo-osmotic
    Beta-lactam treated cells may be saved from lysis by
high salt or sugar

          Backbones of penicillin and D-Ala-D-Ala

Beta-Lactams are analogs of D-Ala-D-Ala
    They prevent cross-linking of peptidoglycan
    C-N bond in lactam ring corresponds to the C-N bond
between the two D-Ala-D-Ala that is broken during cross-

Cross-linking enzyme: a) binds beta-lactam instead
                 b) breaks the lactam (C-N) bond
                 c) attaches beta-lactam to a serine in
active site
                 d) enzyme is now dead
Penicillin Binding Proteins (PBP) of E.
Penicillin binding proteins = enzymes that bind D-Ala-D-Ala

PBP1, 2 and 3 are dual function enzymes
    I) transglycosylases - add new disaccharide units to
         growing chains
    II) transpeptidases - form cross-links between the
         peptide side chains

PBP#    MW (kd) Enzyme Activity            Function
1A      92      transpeptidase             elongation
1B      85       transpeptidase            elongation
2       66       transpeptidase            initiation
3       60       transpeptidase            cross-Wall
4       49       endopeptidase             remodeling
5       42       DD-carboxypeptidase       removes last D-
6       40       DD-carboxypeptidase       removes last D-
7       29       unknown               unknown

Most beta-lactams bind best to PBP-1A and PBP-1B and cause

Mecillinam binds best to PBP-2 and forms giant spherical
    lysis at higher antibiotic concentrations

Cephalexin binds best to PBP-3 and produces filaments with
    lysis at higher antibiotic concentrations

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