and the reference 185 authors hypothesized that the centers could be charac-
terized as two catalytic (C) centers and three electron transfer (E) functional
units. R. L. Lieberman in the Rosenzweig group at Northwestern observed a
type 2 mononuclear Cu(II) EPR signal only. 184b,c,e,186 These workers also pro-
posed a mononuclear copper site plus a copper - containing cluster based on
extended X - ray absorption fi ne structure (EXAFS) best fi tted with a 2.57 - Å
Cu – Cu interaction. 184b,187 In 2005, the researchers were able to crystallize Meth-
ylococcus capsulatus (Bath) pMMO and reported its 2.8 - Å resolution X - ray
crystallographic structure^188 (PDB: 1YEW).
X - ray crystallography of the particulate MMO revealed a trimer composed
of three copies (protomers) each of pmoA, pmoB, and pmoC to form a α 3 β 3 γ 3
cylinder ∼ 105 Å long and ∼ 90 Å in diameter. A soluble region, composed
mainly of sixβ - barrel structures, extends ∼ 45 Å away from the membrane. The
soluble region rests on 42 transmembrane (TM) helices, 14 from each pro-
tomer. In the soluble region, a ∼ 11 - Å hole, lined with hydrophilic residues —
glutamic acid, aspartic acid, and arginines — is located in the center of the
trimer. The hole extends into the TM region, widening eventually to ∼ 22 Å.
The reference 188 authors believe that the trimer is physiologically relevant.
One protomer in the trimer can be described in the following manner using
PDB: 1YEW data as visualized in Figure 7.51. The chain identifi ed as pmoB
includes residues 33 – 414, and it is colored cyan in Figure 7.51. The pmoB sec-
ondary structure includes two antiparallelβ - barrel structures, one at the N -
terminal end and one at the C - terminus. The N - terminal β - barrel includes
seven strands and is oriented∼ 90 ° in relation to the C - terminal β - barrel, which
contains eight strands. The two β - barrels are separated by a β - hairpin followed
by the two TM (transmembrane) helices. The pmoB secondary structural motif
is similar to that of subunit II of cytochrome c oxidase (Section 7.8). Both
pMMO and cytochrome c oxidase contain monomeric copper(II) and dinu-
clear Cu – Cu centers within the β - barrel motif. In cytochrome c oxidase, these
are identifi ed as the dinuclear Cu A and monomeric Cu B sites. In pMMO, the
mononuclear copper ion, colored blue in Figure 7.51 , is located ∼ 25 Å above
the membrane and near the surface of the N - terminal β - barrel. This copper
ion is coordinated by the N δ atoms of his48 and his72, shown in stick fi gure
format in Figure 7.51 , with a nearly linear geometry. Gln404 resides within 3 Å
of this site as well. This mononuclear copper center resembles the type 2 site
found in multicopper oxidases such as ascorbate oxidase and ceruloplasmin,
and it probably is responsible for the type 2 Cu(II) EPR signal seen for puri-
fi ed pMMO. The second pmoB copper ion site, blue in Figure 7.51 , is also
within its N - terminal β - barrel, ∼ 21 Å from the mononuclear site. The site ’ s
location, about 10 Å above the lipid bilayer interface, is similar to that of
cytochrome c oxidase ’ s Cu A site. The Cu – Cu distance in PDB: 1YEW refi nes
to∼ 2.6 Å , similar to the 2.57 Å value found from EXAFS studies. The coordi-
nation sphere for one copper ion includes his33, the N - terminal amino acid
found for the pmoB protomers of PDB: 1YEW. Both the N - terminal amino
nitrogen and the side - chain N δ of his33 are within bonding distance to Cu 1.
NON-HEME IRON-CONTAINING PROTEINS 461