Team:Paris Bettencourt/Project/Eliminate Smell

We analyzed human sweat samples for two things. First, we conducted 16S sequencing of the samples collected in order to determine the types of Corynebacterium species present in the sample. The reason we were interested in Corynebacterium was because it is known from literature that one of the main enzyme responsible for the body odor smell (AgaA) is found in Corynebacterium species. Not only did we do 16S sequencing on these samples, but also conducted smell tests on the same samples in order to see if there was a correlation between odor smell and presence of Corynebacterium species.

[Figure 4: phylogenetic tree] [Figure 5: smell test data??]

Second, we conducted Sanger sequencing of axillary sweat samples for the genes in Figure 1. [MEGANE ADD STUFF HERE ABOUT WHAT YOU DID EXACTLY / HOPED TO DO].

[Figure 6: megane's stuff??]

1. The Microbiome: Looking for Genes Responsible for Body Odor

Introduction
In previous literature, it has been determined that there are a few key enzymes responsible for body odor, such as AgaA, AecD, Ldh, and AckA. Figure 1 shows a description of the enzymes studied in our project and the reactions corresponding to each enzyme.

Figure 1:Enzymes responsible for body odor in the human axilla [ref].

We Sanger-sequenced human axilary sweat samples for these genes.
(MÉGANE'S FIGURE)
(MÉGANE'S RESULTS: XXX genes are responisble for body odor).

The AgaA enzyme in Corynebacterium striatum is found to be a major source of "pungent" or "musky" odor in humans [ref]. A specific bacterial aminoacylase cleaves odorant precursors secreted in the human axilla [ref], and hydrolyzes 3-methyl-2-hexenoyl-glutamine (3M2H-gln) into 3-methyl-2-hexenoic acid (3M2H) and free glutamine. The enzyme is known to have a low specificity for the acyl group, and to act on a range of glutamine conjugates. We cloned agaA into the standard BioBrick vector, and expressed it in E. coli.

In order to analyze the smell created by AgaA, agaA was successfully cloned into E. coli. A noticeable odor was produced by agaA-expressing E. coli grown in selective LB, described as "beer-like" or "cheese-like". We took this to be evidence that the enzyme was functional and acting on a non-native substrate in LB media. We confirmed this observation with a formal smell test (Figure 2) as well as analysis by gas chromatography (Figure 3).



Figure 2:14 people smelled two tubes of E. coli grown to saturation in LB. One culture carried synthetic agaA and other an empty vector control. 13 people out of 14 rated the E. coli carrying agaA as more smelly (pink) than the control(violet). Experiments were double-blind. Significance was confirmed by Chi square test (p-value = 0.001341).

[figure 3 GC analysis]

2) CRISPRs: finding natural odorless mutants

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3) Probiotic cream: a cure for body odor

(Figure for the cream.)